U.S. Environmental Protection Agency
Hazard Characterization Document
December, 2012
SCREENING-LEVEL HAZARD CHARACTERIZATION
Chloronitrobenzenes Category
SPONSORED CHEMICALS
o-Chloronitrobenzene (ONCB) (CASRN 88-73-3)
m-Chloronitrobenzene (MNCB) (CASRN 121-73-3)
p-Chloronitrobenzene (PNCB) (CASRN 100-00-5)
The High Production Volume (HPV) Challenge Program1 was conceived as a voluntary initiative
aimed at developing and making publicly available screening-level health and environmental
effects information on chemicals manufactured in or imported into the United States in quantities
greater than one million pounds per year. In the Challenge Program, producers and importers of
HPV chemicals voluntarily sponsored chemicals; sponsorship entailed the identification and
initial assessment of the adequacy of existing toxicity data/information, conducting new testing if
adequate data did not exist, and making both new and existing data and information available to
the public. Each complete data submission contains data on 18 internationally agreed to "SIDS"
(Screening Information Data Set2) endpoints that are screening-level indicators of potential
hazards (toxicity) for humans or the environment.
The Environmental Protection Agency's Office of Pollution Prevention and Toxics (OPPT) is
evaluating the data submitted in the HPV Challenge Program on approximately 1400 sponsored
chemicals by developing hazard characterizations (HCs). These HCs consist of an evaluation of
the quality and completeness of the data set provided in the Challenge Program submissions.
They are not intended to be definitive statements regarding the possibility of unreasonable risk of
injury to health or the environment.
2 3
The evaluation is performed according to established EPA guidance ' and is based primarily on
hazard data provided by sponsors; however, in preparing the hazard characterization, EPA
considered its own comments and public comments on the original submission as well as the
sponsor's responses to comments and revisions made to the submission. In order to determine
whether any new hazard information was developed since the time of the HPV submission, a
search of the following databases was made from one year prior to the date of the HPV
Challenge submission to the present: (ChemID to locate available data sources including
Medline/PubMed, Toxline, HSDB, IRIS, NTP, AT SDR, IARC, EXTOXNET, EPA SRS, etc.),
STN/CAS online databases (Registry file for locators, ChemAbs for toxicology data, RTECS,
Merck, etc.) and Science Direct and ECHA4. OPPT's focus on these specific sources is based on
their being of high quality, highly relevant to hazard characterization, and publicly available.
1 U.S. EPA. High Production Volume (HPV) Challenge Program; http://www.epa.gov/chemrtk/index.htm.
2 U.S. EPA. HPV Challenge Program - Information Sources; http://www.epa.gov/chemrtk/pubs/general/guidocs.htm.
3 U.S. EPA. Risk Assessment Guidelines; http://cfpub.epa.gov/ncea/raf/rafguid.cfm.
4 European Chemicals Agency, http://echa.europa.eu.
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OPPT does not develop HCs for those HPV chemicals which have already been assessed
internationally through the HPV program of the Organization for Economic Cooperation and
Development (OECD) and for which Screening Initial Data Set (SIDS) Initial Assessment
Reports (SIAR) and SIDS Initial Assessment Profiles (SIAP) are available, but when appropriate
will reference said materials in support of related chemicals. These documents are presented in
an international forum that involves review and endorsement by governmental authorities around
the world. OPPT is an active participant in these meetings and accepts these documents as
reliable screening-level hazard assessments.
These hazard characterizations are technical documents intended to inform subsequent decisions
and actions by OPPT. Accordingly, the documents are not written with the goal of informing the
general public. However, they do provide a vehicle for public access to a concise assessment of
the raw technical data on HPV chemicals and provide information previously not readily
available to the public.
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Chemical Abstract Service
Registry Number
(CASRN)
Sponsored Chemical
88-73-3
121-73-3
100-00-5
Chemical Abstract Index
Name
Sponsored Chemical
Benzene, l-chloro-2-nitro-
Benzene, l-chloro-3-nitro-
Benzene, l-chloro-4-nitro-
Structural Formula
See Section 1 Table 1
Summary
The chloronitrobenzene category consists of three structural isomers: benzene, l-chloro-2-nitro-;
benzene, l-chloro-3-nitro-; and benzene, l-chloro-4-nitro-. All three compounds are yellow
solids with moderate vapor pressure and moderate water solubility. The substances in the
chloronitrobenzene category are expected to possess moderate mobility in soil. Volatilization is
considered moderate based on the Henry's Law constants for these substances. The rate of
hydrolysis is negligible. The rate of atmospheric photooxidation is negligible. The substances in
the chloronitrobenzene category are expected to have moderate persistence (P2) and low
bioaccumulation potential (Bl).
The acute oral, inhalation and dermal toxicity of the chloronitrobenzenes in rats is moderate.
The acute dermal toxicity of PNCB in rabbits is low. In a 13-week, inhalation repeated-dose
toxicity study, rats exposed to ONCB exhibited an increase in methemoglobin levels and
hyperplasia and hypertrophy of the respiratory epithelium at 0.007 mg/L/day; the NOAEC was
not established. In a 13-week, inhalation repeated-dose toxicity study, mice exposed to ONCB
exhibited increased liver and spleen weights and hepatocytomegaly in the liver at 0.0576
mg/L/day; the NOAEC is 0.0288 mg/L/day. In a 5-week dietary study in mice, exposure to
ONCB produced histopathological changes in the liver in males and females at 167 mg/kg-
bw/day and 220 mg/kg-bw/day, respectively; the NOAELmaies is 16 mg/kg-bw/day and the
NOAELfemaies is 24 mg/kg-bw/day. In a 13-week, inhalation repeated-dose toxicity study, mice
exposed to PNCB exhibited organ weight changes and histopathology in the liver at 0.0768
mg/L/day; the NOAEC is 0.0384 mg/L/day. In a 24-month oral gavage study, rats exposed to
PNCB exhibited increased methemoglobin levels at 0.7 mg/kg-day; the NOAEL is 0.1 mg/kg-
day. In an NTP, reproductive assessment by continuous breeding (RACB) study (oral gavage),
mice exposed to ONCB, exhibited an increase in methemoglobin levels and organ weight
changes at 80 mg/kg-day. There were no treatment related effects on the reproductive
parameters examined. The NOAEL for systemic and reproductive toxicity is 160 mg/kg-day
(highest dose tested). In an NTP, reproductive assessment by continuous breeding (RACB)
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study (oral gavage), female mice exposed to PNCB, exhibited decreased fertility and organ
weight changes at 250 mg/kg-day; the NOAEL for systemic and reproductive toxicity was 125
mg/kg-day. In a prenatal developmental toxicity study in rats exposed to ONCB via oral gavage,
pregnant dams exhibited decreased maternal weight gain and increased early resorptions and post
implantation losses at 75 mg/kg-day; the NOAEL for maternal toxicity is 25 mg/kg-day. In a
prenatal developmental toxicity study in rats exposed to PNCB via oral gavage, pregnant dams
exhibited reduced body weight gain and increased spleen weights at 15 mg/kg-day; the NOAEL
for maternal toxicity is 5 mg/kg-day. In the same study, increased resorptions and skeletal
malformations and decreased live births and fetal weight were observed in pups from the 45
mg/kg-day treatment group; the NOAEL for developmental toxicity is 15 mg/kg-day. In the
same study, kits exhibited an increased incidence in fetal malformations at 5 mg/kg-day. In the
two-generation reproductive toxicity study in rats previously described, there was a decrease in
pup survival at 0.1 mg/kg-day; the NOAEL for developmental toxicity was not established.
ONCB and PNCB are mutagenic in bacteria, induced sister chromatid exchange in mammalian
cells and induced chromosomal aberrations in mammalian cells in vitro. MNCB was not
mutagenic in bacteria and did not induce chromosomal aberrations in mammalian cells in vitro.
PNCB did not induce chromosomal aberrations in rats in vivo. PNCB did not induce
unscheduled DNA synthesis in mammalian cells in vitro. ONCB is not irritating to rabbit skin
and is irritating to rabbit eyes. PNCB is irritating to rabbit skin and not irritating to rabbit eyes.
PNCB was carcinogenic to rats in a 24-month oral gavage study.
MNCB (CASRN121-73-3)
The 96-hour LC50 value for fish exposed to MNCB is 18.8 mg/L. The 48-hour EC50 value for
aquatic invertebrates exposed to MNCB is based on read-across from the range of values from
ONCB and PNCB as 2.7 to 41 mg/L. The 96-hour EC50 value for aquatic plants exposed to
MNCB is based on read-across from ONCB and PNCB ranged values of 6.9 to 34 mg/L for
biomass and 4.9 to 16 mg/L for growth rate. The 14-d NOEC for fish exposed to MNCB is based
on read-across from ONCB and PNCB as a range of values from 1.53 to 30.03 mg/L. The 33-day
NOEC for fish exposed to MNCB is based on the read- across value from ONCB as 0.264 mg/L.
The 21-day LOEC for MNCB based on the reproduction endpoint is read-across from PNCB as
0.103. The NOEC for MNCB is based on the range of read-across values from PNCB and ONCB
as 0.190 to 3 mg/L.
ONCB (CASRN 88- 73-3)
The 96-hour LC50 value for fish exposed to ONCB is 25.5 mg/L. The 48-hour EC50 value for
aquatic invertebrates exposed to ONCB ranges from 21.3 to 41 mg/L. The 96 hour EC50 for
aquatic plants exposed to ONCB ranges from 6.9 to 34 mg/L for biomass. The 14-day fish
NOEC for ONCB is 30.03 mg/L. The 33-day NOEC for fish exposed to ONCB is 0.264 mg/L.
The 21-day NOEC for aquatic invertebrates exposed to ONCB is 3 mg/L based on the
reproduction endpoint.
PNCB (CASRN 100-00-5)
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The 96-hour LC50 value for fish exposed to PNCB ranges from 2 to 26.7 mg/L. The 48-hour
EC50 value for aquatic invertebrates exposed to PNCB ranges from 2.7 to 10 mg/L. The 96 hour
EC50 for aquatic plants exposed to PNCB was 8 mg/L for biomass and ranges from 4.9 to 16
mg/L for growth rate. The 14-day NOEC for fish exposed to PNCB is 1.53 mg/L. The 21-day
LOEC and NOEC for aquatic invertebrates exposed to PNCB are 0.103 mg/L and 0.190 mg/L,
respectively, based on the reproduction endpoint.
No data gaps were identified under the HPV Challenge Program.
The sponsor, Solutia, Inc., submitted a Test Plan and Robust Summaries to EPA for the
chloronitrobenzenes category on April 9, 2003. EPA posted the submission on the ChemRTK
HPV Challenge website on April 23, 2003
(http://www.epa.gov/chemrtk/pubs/summaries/chlrnbnz/cl4392tc.htm). EPA comments on the
original submission were posted to the website on August 28, 2003. Public comments were also
received and posted to the website. The sponsor submitted updated/revised documents on June
8, 2004, which were posted to the ChemRTK website on August 27, 2004.
Category Justification
The chloronitrobenzenes category contains three structural isomers of chloronitrobenzene:
ONCB (CASRN 88-73-3), MNCB (CASRN 121-73-3), and PNCB (CASRN 100-00-5). The
chloronitrobenzene isomers were grouped together based on their structural similarity, which
was expected to result in both similar properties and biological modes of action. This
justification was supported by the limited range in toxicity values among category members for
both mammalian and aquatic endpoints. EPA agrees that it is appropriate to group these three
structural isomers together for this screening level hazard characterization based on structural
similarity and physicochemical properties.
1. Chemical Identity
1.1 Identification and Purity
The chloronitrobenzenes are isomeric forms of chloronitrobenzene, where the nitro group is
attached to the chlorobenzene ring at either the ortho, meta, or para position. The purity of the
isomers was > 97% when specified in the Robust Summaries or the TSCATS documents.
The chemical structures and SMILES of the chloronitrobenzenes are depicted in Table 1.
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Table 1. Chloronitrobenzenes Category Sponsored Chemical Structures
Chemical Name
CASRN
Structure
Benzene, l-chloro-2-
nitro-
88-73-3
6r"°'
SMILES: 0=N(=0)c(c(ccc 1 )Cl)c 1
Benzene, l-chloro-3-
nitro-
121-73-3
CI
SMILES: 0=N(=0)c(cccc 1 Cl)c 1
Benzene, l-chloro-4-
nitro-
100-00-5
CI
no2
SMILES: N(=0)(=0)c(ccc(c 1 )Cl)c 1
1.2 Physical-Chemical Properties
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The physical-chemical properties of the substances contained within the chloronitrobenzene
category are summarized in Table 2. The substances contained in the chloronitrobenzene
category are yellow solids with moderate vapor pressure and moderate water solubility.
Table 2. Physical-Chemical Properties of the Chloronitrobenzene Category1
Property
Benzene, l-chloro-2-
nitro-
Benzene, l-chloro-3-
nitro-
Benzene, l-chloro-4-
nitro-
CASRN
88-73-3
121-73-3
100-00-5
Molecular Weight
157.56
157.56
157.56
Physical State
Yellow crystals2
Yellow crystals2
Pale-yellow
orthorhombic prisms
Melting Point
32.5°C (measured)
46°C (measured)
83.4°C (measured)
Boiling Point
245.7°C (measured)
236°C (measured)
242°C (measured)
Vapor Pressure
0.043 mm Hg at 20°C
(measured)
0.097 mm Hg at 25°C
(measured)
0.094 mm Hg at 25°C
(measured)
Dissociation Constant
(pKa)
Not applicable
Not applicable
Not applicable
Henry's Law Constant
9.3xlO"6 atm-m3/mole
"3
(measured)
1.35xlO"5 atm-m3/mole
"3
(measured)
4.89><10"6 atm-m3/mole
"3
(measured)
Water Solubility
198 mg/L at 25°C
(measured)
273 mg/L at 20°C
(measured)
189.4 mg/L at 25°C
(measured)
Log Kow
2.24 (measured)
2.49 (measured)
2.39 (measured)
^olutia Incorporated. 2004. Revised Test Plan and Robust Summary for the
Chloronitrobenzene Category. Available online at
http://www.epa. gov/chemrtk/pubs/summaries/chlrnbnz/c 14392tc.htm as of January 17, 2012.
HSDB. 2012. Hazardous Substance Databank. Available online at
http://toxnet.nlm.nih.gov/cgi-bin/sis/htmlgen7HSDB as of January 17, 2012.
3SRC. The Physical Properties Database (PHYSPROP). Syracuse, NY: Syracuse Research
Corporation. Available from http://www.svrres.com/esc/physprop.htm as of January 17, 2012.
2. General Information on Exposure
2.1 Production Volume and Use Pattern
The Chloronitrobenzenes category contains the following three chemicals. All three chemicals
were not reported in the 2006 IUR.
• CASRN 88-73-3 Benzene, l-Chloro-2-Nitro
• CASRN 121-73-3 Benzene, l-Chloro-3-Nitro
• CASRN 100-00-5 Benzene, l-Chloro-4-Nitro
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2.2 Environmental Exposure and Fate
The environmental fate properties of the substances contained within the chloronitrobenzene
category are summarized in Table 3. The substances in the chloronitrobenzene category are
expected to possess moderate mobility in soil. Ten-month semi-continuous activated sludge
(SCAS) (OECD 302A) test results were available for benzene, l-chloro-2-nitro- and benzene, 1-
chloro-4-nitro. The feeding rate, started at 1 mg/24 hours, was raised in 1 mg increments to 5
mg over 28 days. This rate was maintained at 5 mg/24 hours for 4 months and then raised to 10
mg/24 hours. The inoculum was municipal waste treatment sludge. From days 75-120 (5 mg
feed level, high aeration), 10.6 and 33.9% average disappearance over a 24-hour cycle was
reported for benzene, l-chloro-2-nitro-and benzene, l-chloro-4-nitro, respectively. Over the
next 60 days, (5 mg feed level, high aeration), 37.5 and 30.7% disappearance was reported over a
24-hour cycle for benzene, l-chloro-2-nitro-and benzene, l-chloro-4-nitro, respectively. Over
the last 2 weeks (10 mg feed level, low aeration), average disappearances of 47.7 and 65.1% over
a 24-hour cycle were measured for benzene, l-chloro-2-nitro- and benzene, l-chloro-4-nitro,
respectively. A river die-away test indicated limited degradation for the chloronitrobenzene
category. In this study, benzene, l-chloro-2-nitro- and benzene, l-chloro-4-nitro were added at
an initial concentration of 0.961 mg/L to river water collected from the Mississippi River near St.
Louis, MO. At the end of a 56 day incubation period, the concentration of benzene, l-chloro-2-
nitro- and benzene, l-chloro-4-nitro were 0.904 (5.93% loss) and 0.859 (13.4% loss) mg/L,
respectively. Benzene, l-chloro-2-nitro- was shown to be not readily biodegradable achieving
0%> of its theoretical biochemical oxygen demand (BOD) in the MITI (OECD 301C) and a
closed bottle (OECD 301D) test. Benzene, l-chloro-4-nitro was also not readily biodegradable,
achieving 0% of its theoretical BOD in 14 days in the MITI (OECD 301C) test. These data
suggest that aerobic biodegradation will occur slowly in the environment; however, anaerobic
biodegradation may be an important fate process for the chloronitrobenzene category. Benzene,
l-chloro-3-nitro- had a half-life of approximately 3.2 days using sediment obtained from the
Tsurumi River, Japan and maintained under anaerobic conditions. Volatilization is considered
moderate based on the Henry's Law constants for these substances. The rate of hydrolysis is
negligible. The rate of atmospheric photooxidation is considered negligible. The substances in
the chloronitrobenzene category are expected to have moderate persistence (P2) and low
bioaccumulation potential (Bl).
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Table 3. Environmental Fate Characteristics of the Chloronitrobenzene Category1
Property
Benzene, l-chloro-2-
nitro-
Benzene, l-chloro-3-
nitro-
Benzene, l-chloro-4-
nitro-
CASRN
88-73-3
121-73-3
100-00-5
Photodegradation Half-
life
62 days (estimated)
89 days (estimated)
62 days (estimated)
Hydrolysis Half-life
>1 year
>1 year
>1 year
Biodegradation
11-48% (inherently
biodegradable);
0% in 14 days (not
readily
biodegradable)3;
0% biodegradation in
30 days (not readily
biodegradable)4
Half-life of 3.2 days
(anaerobic sediment)
34-66% (inherently
biodegradable);
0% in 14 days (not
"3
readily biodegradable)
Bioaccumulation
Factor
BCF = 7.0-20.8
(measured in carp at
0.25 ppm)3;
BCF = 7.4-22.3
(measured in carp at
0.025 ppm)3;
BAF = 15.6
(estimated)
BAF = 29.5
(estimated)
BCF = 5.8-20.9
(measured in carp at
0.15 ppm)3;
BCF = 7.5-18.1
(measured in carp at
0.015 ppm)3;
BAF = 21.7
(estimated)
Log Koc
2.6 (estimated)2
2.6 (estimated)2
2.6 (estimated)2
Fugacity
(Level III Model)2
Air (%)
Water (%)
Soil (%)
Sediment (%)
3.1
19.1
77.5
0.4
3.2
18.9
77.5
0.4
2.8
19.2
77.6
0.4
Persistence5
P2 (moderate)
P2 (moderate)
P2 (moderate)
Bioaccumulation5
B1 (low)
B1 (low)
B1 (low)
^olutia Incorporated. 2004. Revised Test Plan and Robust Summary for the
Chloronitrobenzene Category. Available online at
http://www.epa. gov/chemrtk/pubs/summaries/chlrnbnz/c 14392tc.htm as of January 17, 2012.
2U.S. EPA. 2012. Estimation Programs Interface Suite™ for Microsoft® Windows, v4.10.
U.S. Environmental Protection Agency, Washington, DC, USA. Available online at
http://www.epa. gov/opptintr/exposure/pubs/episuitedl.htm as of January 17, 2012.
"3
National Institute of Technology and Evaluation. 2002. Biodegradation and Bioaccumulation
of the Existing Chemical Substances under the Chemical Substances Control Law. Available
online at http://www.safe.nite.go.ip/english/kizon/KIZON start hazkizon.html as of January
17, 2012.
4Kaiser KLE. 1998. Review of biodegradability tests for the purpose of developing regulations.
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Table 3. Environmental Fate Characteristics of the Chloronitrobenzene Category1
Property
Benzene, l-chloro-2-
nitro-
Benzene, l-chloro-3-
nitro-
Benzene, l-chloro-4-
nitro-
Water Qual Res J Canada 33:185-211.
5Federal Register. 1999. Category for Persistent, Bioaccumulative, and Toxic New Chemical
Substances. Federal Register 64, Number 213 (November 4, 1999) pp. 60194-60204.
Conclusion: The chloronitrobenzene category consists of three structural isomers: benzene, 1-
chloro-2-nitro-; benzene, l-chloro-3-nitro-; and benzene, l-chloro-4-nitro-. All three compounds
are yellow solids with moderate vapor pressure and moderate water solubility. The substances in
the chloronitrobenzene category are expected to possess moderate mobility in soil.
Volatilization is considered moderate based on the Henry's Law constants for these substances.
The rate of hydrolysis is negligible. The rate of atmospheric photooxidation is negligible. The
substances in the chloronitrobenzene category are expected to have moderate persistence (P2)
and low bioaccumulation potential (Bl).
3. Human Health Hazard
A complete OECD SIDS dossier on ONCB (CASRN 88-73-3) can be found at:
http ://www. chem .unep. ch/irptc/sids/OECD SID S/CHLORONITROB .pdf
A complete OECD SIDS dossier on PNCB (CASRN 100-00-5) can be found at:
http://www.chem.unep.ch/irptc/sids/QECDSIDS/100005.pdf
A summary of health effects data submitted for SIDS endpoints is provided in Table 4. The table
also indicates where data for tested category members are read-across (RA) to untested members
of the category.
Acute Oral Toxicity
ONCB (CASRN 88- 73-3)
Summaries 1-5 can be found in the complete OECD SIDS dossier on ONCB found at:
http://www.chem.unep.ch/irptc/sids/OECDSIDS/CHLORONITROB.pdf
(1) Male rats (15/dose; strain not specified) were administered ONCB in polyethylene glycol
400 via gavage at 50, 100, 150, 200, 250, 300, or 500 mg/kg and observed for 14 days. Mortality
rates were 0/15, 0/15, 2/15, 4/15, 10/15, 14/15, and 15/15 at 50, 100, 150, 200, 250, 300, and 500
mg/kg, respectively.
LD50= 219 mg/kg
(2) Female rats (15/dose; strain not specified) were administered ONCB in polyethylene glycol
400 via gavage at 25, 50, 100, 250, 350, 500, 650, or 850 mg/kg and observed for 14 days.
Mortality rates were 0/15, 0/15, 015, 1/15, 2/15, 10/15, 12/15, and 15/15 at, 50, 100, 250, 350,
500, 650, or 850 mg/kg, respectively.
LD50 = 457 mg/kg
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(3) Male Wistar rats (10/dose) were administered ONCB in Lutrol via gavage at 100, 200, 250,
300, or 400 mg/kg and observed for 14 days. Mortality rates were 0/10, 2/10, 5/10, 7/10, and
10/10 at 100, 200, 250, 300, or 400 mg/kg, respectively.
LD5o = 251 mg/kg
(4) Female Wistar rats (10/dose) were administered ONCB in Lutrol via gavage at 100, 200, 300,
400, or 500 mg/kg and observed for 14 days. Mortality rates were 0/10, 3/10, 5/10, 9/10, and
10/10 at 100, 200, 300, 400, or 500 mg/kg, respectively.
LD50 = 263 mg/kg
(5) Sprague-Dawley rats (5 mixed sex/dose) were administered ONCB in corn oil via gavage at
398, 501, 631 or 794 mg/kg and observed for 7 days following dosing. Mortality rates were 1/5,
2/5, 4/5 and 5/5 at 398, 501, 631, or 794 mg/kg, respectively.
LD50 = 560 mg/kg
MNCB (CASRN121-73-3)
Sprague-Dawley rats (5/sex/dose) were administered MNCB via gavage at 0, 200, 251, 316, 398
or 501 mg/kg and observed for 14 days following dosing. Mortalities were observed at 251, 316,
398 and 501 mg/kg.
LD50 = 400 mg/kg
PNCB (CASRN 100-00-5)
(1) Sprague-Dawley rats (5 mixed sex/dose) were administered PNCB via gavage at 398, 501,
631, or 794 mg/kg in 10% corn oil and observed for 14 days. Mortality rates were 1/5, 2/5, 4/5,
and 5/5 at dose levels of 398, 501, 631, and 794 mg/kg, respectively.
LD50 = 530 mg/kg
(2) Male Wistar rats (10/dose) were administered PNCB in polyethylene glycol via gavage at
100, 200, 300, 350, 400, 500, or 600 mg/kg and observed for 14 days following dosing.
Mortality rates were 0/10, 3/10, 5/10, 5/10, 8/20, 8/10, and 10/10 at 100, 200, 300, 350, 400, 500,
and 600 mg/kg, respectively.
LD50 = 294 mg/kg
Acute Inhalation Toxicity
ONCB (CASRN 88- 73-3)
Crl:CD rats (10 males/dose) were exposed head-only to ONCB as a vapor/aerosol at 1.56, 1.83,
2.46, 2.64, 3.23 or 3.33 mg/L for 4 hours and observed for 14 days following exposure.
Mortality rates were 1/10, 3/10, 2/10, 10/10, 1/10, and 6/10 at 1.56, 1.83, 2.46, 2.64, 3.23 and
3.33 mg/L, respectively. Additional details are from TSCATS (OTS0540655).
LC50 = 3.20 mg/L
PNCB (CASRN 100-00-5)
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Sprague-Dawley rats (10 males/dose) were exposed head-only to PNCB as a vapor/aerosol at
2.63, 2.84, 3.27, 3.35, 3.73, 6.12, 9.47 or 16.1 mg/L for 4 hours and observed for 14 days
following exposure. One mortality was observed at the highest dose. Additional details are from
TSCATS submission (OTS0557112).
LC50 > 16.1 mg/L
Acute Dermal Toxicity
ONCB (CASRN 88- 73-3)
Summaries 2 and 3 can be found in the complete OECD SIDS dossier on ONCB found at:
http://www.chem.unep.ch/irptc/sids/OECDSIDS/CHLORONITROB.pdf
(1) New Zealand Albino rabbits (2/sex/dose) were administered ONCB (undiluted) via the
dermal route at 251, 316, 398, 501 or 631 mg/kg under unspecified conditions for a single 24-
hour exposure and observed for 14 days following application. Male mortalities were observed
at doses >501 mg/kg and female mortalities were observed at doses >316 mg/kg. Additional
details are from TSCATS (OTS0546300).
LD50 = 400 mg/kg
(2) Male Wistar rats (10/dose) were administered ONCB in polyethylene glycol 400 via the
dermal route at 250, 350, 500, 750, 1000, or 1500 mg/kg, on shaved skin under occluded
conditions for 24 hours and observed for 14 days. Mortality rates were 1/10, 1/10, 3/10, 7/10,
7/10, and 9/10 at 250, 350, 500, 750, 1000, or 1500 mg/kg.
LD50 = 655 mg/kg
(3) Female Wistar rats (10 or 20 rats/dose) were administered ONCB in polyethylene glycol 400
via the dermal route at 750, 1000, or 1500 mg/kg, on shaved skin under occluded conditions for
24 hours and observed for 14 days. Mortality rates were 0/10, 5/10, and 6/10 at 750, 1000, and
1500 mg/kg.
LD50 = 1320 mg/kg
PNCB (CASRN 100-00-5)
(1) Male Wistar rats (10/dose) were administered PNCB in polyethylene glycol via the dermal
route at 500, 600, 700, 900, 1000, or 1200 mg/kg for 24 hours with occlusion and observed for
14 days. Mortality rates were 0/10, 2/10, 5/10, 8/10, 8/10, and 10/10 at 500, 600, 700, 900,
1000, and 1200 mg/kg, respectively.
LD50 = 750 mg/kg
(2) New Zealand Albino rabbits (2/sex/dose) were administered PNCB (undiluted) via the
dermal route on intact skin at 2000, 2510, 3160, 3980 or 5010 mg/kg as a single 24-hour
exposure and observed for 14 days following application. Male and female mortalities were
observed at 3160, 3980 and 5010 mg/kg. Female mortalities were also observed at 2510 mg/kg.
Additional details are from TSCATS (OTS0557080).
LD50 = 3020 mg/kg
Repeated-Dose Toxicity
ONCB (CASRN 88- 73-3)
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(1) In an NTP study, Fischer 344/N rats (10/sex/dose) were exposed to ONCB via inhalation
(whole-body), as a vapor at 0, 7, 14.7, 28.8, 57.6 or 115.2 mg/m3/day (0, 0.007, 0.0147, 0.0288,
0.0576 and 0.1152 mg/L/day) for 6 hours/day, 5 days/week, for 13 weeks. Animals were
evaluated for clinical chemistry, hematology, histopathology and reproductive system effects.
No mortalities were observed. Significant increases in clinical chemistry parameters including:
albumin, total protein concentrations, alanine aminotransferase and sorbitol dehydrogenase were
observed in both sexes at > 57.6 mg/m3/day. Methemoglobinemia was observed and resulted in
normocytic, normochromic anemia with evidence of a hematopoietic response by the end of the
study. Methemoglobin concentrations were significantly increased in males and females treated
"3
with > 7 mg/m /day. Other treatment-related effects on hematological parameters included
decreases in hematocrit, hemoglobin concentrations and erythrocyte counts in exposed males and
3 3
females at > 14.7 mg/m /day and increased reticulocytes in males at > 57.6 mg/m /day and in
females at > 28.8 mg/m3/day. Treatment-related effects included significantly increased liver
3 3
weights for males exposed to > 14.7 mg/m /day and females exposed to > 28.8 mg/m /day.
Significantly increased absolute and relative spleen and lung weights were observed in males
exposed to 115.2 mg/m3/day. Females exposed to > 28.8 mg/m3/day exhibited significantly
increased absolute and relative spleen weights. Treatment-related observations at necropsy were
limited to dark spleens in 2/10 males and 1/10 females in the 115.2 mg/m3/day treatment group.
Treatment-related microscopic observations in the liver included cytoplasmic basophilia of
centrilobular hepatocytes in males and females exposed to > 57.6 mg/m3/day. Treatment-related
microscopic observations in the kidney included cytoplasmic pigment within proximal
convoluted tubule cells in males exposed to > 28.8 mg/m3/day and females exposed to > 57.6
"3
mg/m /day, as well as a concentration-dependent increase in the incidence and severity of tubule
regeneration in males. Hyperplasia/hypertrophy of the respiratory epithelium was a treatment-
related effect observed in the nasal cavity. Treatment-related effects on reproductive parameters
were limited to significantly lower left cauda epididymal weight and significantly lower
"3
spermatid heads per testis and spermatid count of males exposed to 115.2 mg/m /day. See
human health data at http://www.chem.unep.ch/irptc/sids/OECDSIDS/CHLORONITROB. pdf.
LOAEC = 0.007 mg/L/day (based on increased methemoglobin and histopathological changes
in the respiratory epithelium)
NOAEC = Not established
(2) In an NTP study, B6C3F1 mice (10/sex/dose) were exposed to ONCB via inhalation (whole-
body), as a vapor at 0, 7, 14.7, 28.8, 57.6 or 115.2 mg/m3/day (0, 0.007, 0.0147, 0.0288, 0.0576
and 0.1152 mg/L/day) for 6 hours/day, 5 days/week, for 13 weeks. Animals were evaluated for
histopathology and reproductive system effects. Mortalities observed during the study were
limited to two males in the 115.2 mg/m3/day group. The livers of the decedents were
characterized by diffuse, severe sinusoidal congestion with hepatocellular degeneration and
necrosis. Treatment-related effects included increased absolute and relative right kidney weights
"3
and relative liver weights in males exposed to > 14.7 mg/m /day and increased absolute liver
weights in males exposed to >57.6 mg/m3/day. Absolute liver weights in all groups of exposed
"3
females, relative liver weights in females exposed to > 57.6 mg/m /day and absolute right kidney
weights in females exposed to > 14.7 mg/m3/day were significantly increased. Treatment-related
gross observations at necropsy included pale discoloration of the liver in males and one female
exposed to 115.2 mg/m3/day and an enlarged spleen in females exposed to > 57.6 mg/m3/day.
"3
Treatment-related effects on the liver included hepatocytomegaly at > 57.6 mg/m /day and
hepatocellular necrosis, mineralization, and chronic inflammation at 115.2 mg/m3/day .
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Increased hematopoietic activity in the red pulp of the spleen was a minimal treatment effect on
"3
both sexes of mice, particularly in females in the 115.2 mg/m /day group. Sperm morphology
and vaginal cytology evaluations were performed on mice exposed to ONCB at 0, 28.8, 57.6 or
"3
115.2 mg/m /day. Significantly decreased sperm motility was observed in all three treatment
groups. No significant changes in vaginal cytology were observed in females exposed to > 28.8
"3
mg/m /day. See human health data at
http ://www. chem .unep. ch/irptc/sids/OECD SID S/CHLORONITROB .pdf.
LOAEC = 0.0576 mg/L/day (based on organ weight changes and histopathological changes in
the liver)
NOAEC = 0.0288 mg/L/day
(3) Sprague-Dawley rats (15/sex/group) were exposed to ONCB via inhalation (whole-body) at
0, 10, 30 or 60 mg/m3/day (0, 0.01, 0.03 and 0.06 mg/L/day, respectively) for 6 hours/day, 5
days/week, for 4 weeks. Analytical values of test concentrations were 0, 9.9, 30 and 59
mg/m3/day. Parameters monitored in this study included daily morbidity and mortality checks,
weekly detailed clinical observations and body weights, hematology, clinical chemistry and
ophthalmology. Microscopic examination of over 40 tissues and organs was performed on
10 rats/sex from the high-dose and controls and spleens from 10 male and 10 female mid- and
low-dose animals. No effects were seen in ocular toxicity, body weight gain or clinical signs and
no deaths occurred. Animals exhibited an increase in methemoglobin levels at all tested
concentrations; these changes were statistically significant as compared to controls at > 30
"3
mg/m . In female rats, RBCs, hemoglobin and hematocrit levels were significantly decreased as
compared to controls at > 30 mg/m3 and reticulocytes were significantly increased at 60 mg/m3.
In male rats, RBCs, hemoglobin and hematocrit levels were significantly decreased as compared
to controls at 60 mg/m3 and reticulocytes were significantly increased at 60 mg/m3. Male rats
"3
exhibited statistically significant increases in relative liver weight at > 10 mg/m and absolute
liver weight and absolute and relative kidney weight at > 30 mg/m3 while females exhibited
statistically significant increases in absolute and relative liver weight and relative kidney weight
at > 30 mg/m3. All animals exhibited statistically significant increases in absolute and relative
"3
spleen weight at > 30 mg/m . Microscopic findings included increased hemosiderin staining in
the spleen of all animals with an increase in intensity with increasing concentration and an
"3
increase in the degree of extramedullary hematopoiesis at > 30 mg/m in both males and females
(Nair et al., 1986). See human health data at
http ://www. chem .unep. ch/irptc/sids/OECD SID S/CHLORONITROB .pdf.
LOAEC = 0.01 mg/L/day (based on increased methemoglobin and histopathological changes in
the spleen)
NOAEC = Not established
(4) B6C3F1 mice (12/sex/group and an additional 6/sex/group for interim sacrifice) were
administered ONCB via the diet to at doses of 0, 50, 500, or 5000 ppm (0, 16, 167, or 1120
mg/kg-bw in males and 0, 24, 220, or 1310 mg/kg-bw in females) for 5 weeks. One male animal
died in the low-dose group; no other mortalities were reported. Reduced body weight gain,
increased spleen weight, discolored spleen, deposition of hemosiderin in the spleen and increased
liver weight were observed in animals at 5000 ppm. Hematologic changes were observed in
males and females at 5000 ppm, including: reduced erythrocyte count, changes in RBC
morphology, reduced hematocrit and hemoglobin and increased methemoglobin, mean cell
volume of RBCs (MCV), mean cell hemoglobin (MCH), mean cell hemoglobin concentration
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(MCHC), and bilirubin. Changes in enzyme activity were also observed in males at 5000 ppm
and in females at unspecified doses. Centrilobular hepatocytomegaly was observed upon
histopathological examination in males and females at > 500 ppm. Males exhibited decreased
testes weight at 5000 ppm; there were no histopathological changes in the testes. See human
health data at http ://www. chem .unep. ch/irptc/sids/OECD SID S/CHLORONITROB. pdf.
LOAELmaies = 167 mg/kg-bw/day (based on histopathological changes in the liver)
LOAELfemaies = 220 mg/kg-bw/day (based on histopathological changes in the liver)
NOAELmaies = 16 mg/kg-bw/day
NOAELfemaies = 24 mg/kg-bw/day
PNCB (CASRN100-00-5)
(1) In an NTP study, Fischer 344/N rats (10/sex/dose) were exposed to PNCB as a vapor (whole-
body) at 0, 9.6, 19.2, 38.4, 76.8 or 153.6 mg/m3/day (0, 0.0096, 0.0192, 0.0384, 0.0768 and
0.1536 mg/L/day) for 6 hours/day, 5 days/week, for 13 weeks. Animals were evaluated for
histopathology, clinical chemistry, hematology and reproductive system effects. Treatment-
related effects on hematological parameters included methemoglobinemia and a responsive
macrocytic, hyperchromic anemia. Methemoglobin concentrations were significantly increased
in males and females treated with > 9.6 mg/m /day. Decreases in hematocrit, hemoglobin
concentrations and erythrocyte counts were observed in all groups of exposed male and female
rats. Mean corpuscular volume, mean corpuscular hemoglobin and mean cell hemoglobin
"3
concentrations were consistently increased in males and females exposed to > 76.8 mg/m /day.
Reticulocyte counts were increased for male and female rats exposed to > 19.2 mg/m3/day.
Increased numbers of nucleated RBCs accompanied the increases in reticulocyte counts in
exposed males and females. Treatment-related effects with respect to clinical chemistry
parameters included decreased globulin and/or total protein concentrations in male and female
rats exposed to > 38.4 mg/m3/day, increased sorbitol dehydrogenase activity, elevations in bile
"3
acid concentrations in males exposed to > 19.2 mg/m /day and in females exposed to > 76.8
mg/m3/day and decreased serum alkaline phosphatase activity in males exposed to >76.8
3 3
mg/m /day and females exposed to > 38.4 mg/m /day. Treatment-related effects on organ
weights included increased absolute and relative spleen weights in males exposed to > 19.2
3 3
mg/m /day and females exposed to > 76.8 mg/m /day and increased liver weights in females
exposed to > 76.8 mg/m3/day and males exposed to 153.6 mg/m3/day. Increased relative heart
weight and absolute and relative kidney, liver and thymus weights, in addition to decreased
absolute and relative right testis weights, were observed in males exposed to 153.6 mg/m3/day.
Absolute and relative heart and thymus weights were slightly increased in females exposed to >
76.8 mg/m3/day. Significantly increased absolute and relative right kidney weights were
"3
observed in females exposed to 153.6 mg/m /day. Treatment-related effects observed at
necropsy included a concentration-related increased incidence of enlarged or enlarged and
darkened spleens in male and female rats. Treatment-related microscopic observations of the
spleen included congestion of red pulp in all exposed rats of each sex, increased hemosiderin
pigment in all treated animals, increased incidence of hematopoietic cell proliferation in animals
exposed to > 19.2 mg/m3/day and a concentration-dependent increase in incidence and severity
of capsular fibrosis. Treatment-related effects on the kidney included hyaline droplet
nephropathy in males and accumulation of brown pigment granules in males exposed to
3 3
>76.8 mg/m /day and females exposed to > 38.4 mg/m /day. The only observed treatment-
related effect in the liver was increased pigment in the Kupffer cells for males exposed to
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3 3
>76.8 mg/m /day and for females exposed to > 19.2 mg/m /day. Treatment-induced
hematopoietic cell proliferation was observed in the bone marrow of male and female rats
exposed to > 19.2 mg/m3/day. Enlarged mediastinal lymph nodes observed at necropsy
corresponded microscopically to histiocytic hyperplasia in male and female rats exposed to
>76.8 mg/m3/day. Treatment-related effects observed on reproductive parameters included
moderate testicular atrophy characterized by decreased cellularity of seminiferous tubules, lower
left epididymal weights, lower cauda epididymal weights, lower testis weights, lower number of
spermatid heads per testis, lower spermatid count and lower spermatozoal concentration in all
male rats exposed to 153.6 mg/m3/day. Other treatment-induced observations included higher
incidences of infiltrates of chronic inflammatory cells in the harderian gland for male and female
rats exposed to 153.6 mg/m3/day. See human health data at
http://www.chem.unep.ch/irptc/sids/QECDSIDS/100005.pdf
LOAEC = 0.0096 mg/L/day (based on increased methemoglobin and histopathological changes
in the spleen)
NOAEC = Not established.
(2) In an NTP study, B63Flmice (10/sex/dose) were exposed to PNCB as a vapor (whole-body)
at 0, 9.6, 19.2, 38.4, 76.8 or 153.6 mg/m3/day (0, 0.0096, 0.0192, 0.0384, 0.0768 and 0.1536
mg/L/day) for 6 hours/day, 5 days/week, for 13 weeks. Animals were evaluated for
"3
histopathology and reproductive system effects. One male in the 38.4 mg/m /day exposure
group died during the study; the mortality was not clearly related to exposure to PNCB.
Treatment-related effects on organ weights included increased spleen weights of males and
females exposed to > 76.8 mg/m3/day, concentration-related mildly increased relative liver
weights in males and females and significantly increased absolute right kidney weights of all
groups of exposed males and of females exposed to > 19.2 mg/m3/day. Treatment-related gross
lesions consisted of enlarged and dark spleens in male and female mice exposed to 153.6
mg/m3/day and female mice exposed to 76.8 mg/m3/day. Splenic lesions observed
microscopically included minimal to mild congestion, increased hematopoietic cell proliferation
and hemosiderin in animals exposed to > 76.8 mg/m3/day. These observations were present in
"3
all mice exposed to 153.6 mg/m /day. Increased hematopoietic activity and hemosiderin were
observed in most mice exposed to 76.8 mg/m3/day. Increased hematopoiesis and hemosiderin
deposition were also treatment-related effects in the bone marrow of mice exposed to 153.6
mg/m3/day. Hemosiderin deposition in Kupffer cells was the most common finding in the liver
"3
and was limited to male and female mice in the 153.6 mg/m /day groups. Additional treatment-
related changes in the liver included single cell necrosis and hyperplasia in males exposed to
"3
>76.8 mg/m /day and centrilobular cytoplasmic basophilia of hepatocytes in males exposed to
153.6 mg/m3/day. Squamous cell hyperplasia of the forestomach epithelium was observed in
"3
female mice of the 153.6 mg/m /day exposure group. Treatment-related effects on reproductive
parameters included significantly increased estrus cycle length in females exposed to 153.6
"3
mg/m /day. See human health data at
http://www.chem.unep.ch/irptc/sids/QECDSIDS/100005.pdf
LOAEC = 0.0768 mg/L/day (based on increased organ weights and histopathology in the liver)
NOAEC = 0.0384 mg/L/day
(3) Sprague-Dawley rats (10/sex/dose) were exposed to PNCB (> 99% pure) as an aerosol
"3
(whole-body) at mean analytical concentrations of 0, 5, 15 or 45 mg/m /day (0, 0.005, 0.015 or
0.045 mg/L/day) for 6 hours/day, 5 days/week for 4 weeks. No mortalities were observed in
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treated groups and mean body weights of treated animals were similar to control values.
Cyanosis was observed in both males and females and found to be concentration-dependent.
Significant hematologic changes include: dose-dependent increases in methemoglobin in males
3 3 3
at > 5 mg/m /day and in females at > 15 mg/m /day, decreased erythrocytes at > 15 mg/m /day,
decreased hemoglobin at 45 mg/m3/day and decreased hematocrit at 45 mg/m3/day in males and
"3
at > 5 mg/m /day in females. In males, absolute and relative liver and spleen weights were
increased at 45 mg/m3/day. In females, relative liver weights and absolute and relative spleen
"3
weights were increased at 45 mg/m /day. Histopathological changes included, increases in
splenic congestions, extramedullary hematopoiesis and hemosiderosis in the spleen at 5 and 15
"3
mg/m /day.
LOAEC = 0.005 mg/L/day (based on increased methemoglobin and histopathological changes
in the spleen)
NOAEC = Not established
(4) Sprague-Dawley rats (20/sex/dose) were administered PNCB in corn oil via gavage at 0, 3,
10 or 30 mg/kg for 90 days. Blood and urine samples were taken twice during the study to
evaluate hematology, urinalysis and blood chemistry parameters. Microscopic evaluation was
performed on the following tissues: aorta, brain, eye, heart, pituitary, adrenals, trachea, stomach,
skin, pancreas, large intestine, small intestine, kidneys, liver, lung, mesenteric lymph node,
spleen, muscle, prostate/uterus, bone marrow, bone, testis, ovary, thyroids, urinary bladder and
lesions or abnormal masses. No treatment-related effects were observed with respect to body
weights. Statistically significant increases in food consumption were observed in animals treated
with >10 mg/kg. Treatment-related clinical observations included general paleness immediately
after dosing in females treated with >10 mg/kg and in males treated with 30 mg/kg. Treatment-
related effects on hematology at 7 weeks included a significant dose-related decrease in numbers
of circulating RBCs, hemoglobin levels and hematocrit in males treated with > 3 mg/kg and
females treated with >10 mg/kg. These changes were accompanied by dose-related,
significantly increased mean cell volumes (MCV) and mean cell hemoglobin (MCH) for animals
treated with >10 mg/kg. Dose-related, significant decreases in mean cell hemoglobin
concentration (MCHC) were also observed at > 10 mg/kg. These observations were repeated at
13 weeks at all daily doses. Treatment-related effects on urinalysis parameters were limited to
increased levels of urobilinogen in all of the male and female test groups at week 13. Effects on
serum chemistry included a significant decrease in total serum protein for females treated with >
10 mg/kg at week 7 and males treated with >10 mg/kg at week 13, as well as remarkably lower
serum glutamate pyruvate transaminase (SGPT) levels in males treated with 30 mg/kg at week
13. Dose-dependent, significantly increased methemoglobin levels were observed in all test
groups at week 7 and 13. Dose-dependent, significantly increased reticulocyte counts were
observed at the end of the study at all daily dosages, with females having slightly higher counts
than males. Treatment-related effects on gross pathology observed in both sexes included
abnormal coloration of the spleen at all dosages and discoloration of the kidneys in animals
treated with 30 mg/kg. Observations in females included significant enlargement of the heart
and liver at 30 mg/kg and significant enlargement of the spleen at all daily dosages. Gross
pathology observations specific to males included significantly enlarged livers at 30 mg/kg,
kidneys at > 10 mg/kg and spleen at all levels. Treatment-related effects with respect to
histopathology included the presence of excessive hemosiderin in the spleen, excessive
hemopoiesis in the spleen and liver, congestion and vacuolization of congested red pulp in the
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spleen, hemosiderosis in livers and kidney tubules and hyperplasia of the bone marrow (30
mg/kg only). The incidence and severity of histopathological findings were dose-dependent and
one or more of the effects were observed in animals in the 3 mg/kg dose group. Additional
details are from TSCATS (OTS0538501). See human health data at
http://www.chem.unep.ch/irptc/sids/QECDSIDS/100005.pdf
LOAEL = 3 mg/kg-day (based on increased methemoglobin, hematologic changes and organ
weight changes)
NOAEL = Not established
(5) Sprague-Dawley rats (60/sex/dose) were administered PNCB via gavage in corn oil at 0.1,
0.7 or 5.0 mg/kg-day for > 24 months. Physical observations, body weight and food
consumption measurements were performed pretest and at selected intervals throughout the
study. Hematology, clinical chemistry parameters and urinalyses were evaluated at 6, 12, 18 and
24 months. All animals were subject to gross post-mortem examinations and histopathological
evaluation of selected tissues. Tissues examined in the control and 5.0 mg/kg-day-dose groups
included: adrenals, bone and bone marrow, brain, epididymis, esophagus, eye, gonad, heart,
intestine, kidney, liver, lungs, lymph nodes, mammary gland, right sciatic nerve, pancreas,
parathyroid, pituitary, prostate, salivary gland, seminal vesicles, skeletal muscle, skin, spinal
cord, spleen, stomach, thymus, thyroid, trachea, urinary bladder, uterus, gross lesions and tissue
masses. Gross lesions: testes, epididymis and spleens were examined in animals treated with 0.1
and 0.7 mg/kg-day. Weighed organs included adrenals, brain, liver, ovaries, heart, spleen,
kidneys and testes. No treatment-related effects were observed with respect to mortality,
physical observations, body weights, food consumption, ophthalmoscopic examinations, total
neoplasm incidence, clinical chemistry studies and urinalyses. Some observed mortalities (in
control and treated groups) were attributed to intubation accidents and mortality data were
considered comparable between treated and control animals. Observed effects of PNCB
administration included a dose-related increase in methemoglobin in animals treated with >0.7
mg/kg-day. Slight anemia was observed in males and females at 5.0 mg/kg-day; this effect was
demonstrated by slightly decreased hemoglobin, hematocrit and erythrocyte values with
concomitant slight increases in numbers of reticulocytes. Treatment-related effects revealed
upon microscopic examination included elevated spleen weights as well as increased incidence
and/or severity of accumulation of brown pigment (hemosiderin) in the spleens of males and
females treated with 5.0 mg/kg-day. A higher incidence of interstitial cell tumors of the testes in
treated animals was observed compared to control animals. The authors believe this is due to an
unusually low incidence of these tumors in the control group rather than an oncogenic effect of
PNCB. Additional details are from TSCATS (OTS0557088).
LOAEL = 0.7 mg/kg-day (based on increased methemoglobin)
NOAEL = 0.1 mg/kg-day
Reproductive Toxicity
ONCB (CASRN 88- 73-3)
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(1) In the 13-week repeated-dose inhalation study in Fisher 344/N rats described above, sperm
morphology and vaginal cytology evaluations were performed on rats exposed to ONCB at 0,
28.8, 57.6 or 115.2 mg/m /day (animals from the 7 and 14.7 mg/m3/day groups were not
evaluated). Treatment-related effects on reproductive parameters were limited to significantly
lower left cauda epididymal weight and significantly lower spermatid heads per testis and
"3
spermatid count of males exposed to 115.2 mg/m /day. See human health data at
http ://www. chem .unep. ch/irptc/sids/OECD SID S/CHLORONITROB .pdf.
(2) In the 13-week repeated-dose inhalation study in B6C3F1 mice described above, sperm
morphology and vaginal cytology evaluations were performed on mice exposed to ONCB at 0,
28.8, 57.6 or 115.2 mg/m /day (animals from the 7 and 14.7 mg/m3/day groups were not
evaluated). Significantly decreased sperm motility was observed in all three treatment groups.
No significant changes in vaginal cytology were observed in females exposed to > 28.8
"3
mg/m /day. See human health data at
http ://www. chem .unep. ch/irptc/sids/OECD SID S/CHLORONITROB .pdf.
(3) In an NTP, reproductive assessment by continuous breeding (RACB) study, Swiss CD-I
mice (20/sex/dose, treated; 40 pairs, controls) were administered ONCB in corn oil, via gavage
at 0, 40, 80, or 160 mg/kg-day from 1 week prior to cohabitation, through 14 weeks of
cohabitation for the F0 generation. The dams were allowed to nurse the F1 generation and these
animals were weaned at 21 days. The F1 generation was then exposed to the test substance at 0
or 250 mg/kg from 1 week prior to cohabitation, through 14 weeks of cohabitation. For the F0
generation, there were no treatment-related mortalities. Three females from the 160 mg/kg-day
group appeared cyanotic. No other changes in clinical signs were observed. No treatment-
related effects on reproductive function in F0 mice were observed. There were no effects on the
number of litters, pup weight or viability. Pup body weight at weaning was decreased (10-13%)
at > 40 mg/kg-day. After weaning, 12 F0 mice from the control and 160 mg/kg-day treatment
groups were killed and examined for methemoglobin levels and spleen weights. Relative spleen
weight was increased (50-100%) and methemoglobin levels were increased 4- to 6-fold. No
other necropsy data were collected. From the F1 mating, no effects were observed on the
number of litters per group, the number of live pups per litter, pup weight or viability. At
necropsy of the F1 animals, liver and spleen weights were increased in males (40 and 60%,
respectively) and females (40% each) at 250 mg/kg-day. In male mice, absolute right epididymis
and kidney/adrenals weights were increased and relative seminal vesicle weight was slightly
reduced (7%); no other adverse effects on sperm were observed. There were no effects on
estrous cycle length compared to controls. Methemoglobin was increased 3-fold in animals
treated with 250 mg/kg-day (Chapin and Sloane, 1996; Chapin, 1997a). See human health data
at http://www.chem.unep.ch/irptc/sids/OECDSIDS/CIILORONITROB.pdf.
NOAEL (systemic toxicity) = 160 mg/kg-day (highest dose tested)
NOAEL (reproductive toxicity) = 160 mg/kg-day (highest dose tested)
PNCB (CASRN100-00-5)
(1) In the 13-week repeated-dose inhalation study in Fisher 344/N rats described above,
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sperm morphology and vaginal cytology evaluations were performed on rats exposed to PNCB at
0, 6, 12, and 24 ppm. Females in all groups exhibited decreased estrus cycle length. The
remainder of the observed treatment-related effects were limited to males treated with 24 ppm
"3
(153.6 mg/m /day), and included significantly lower left epididymal, cauda epididymal and testis
weights and lower spermatid heads/testis, spermatid counts and spermatozoa concentrations than
control rats. See human health data at
http://www.chem.unep.ch/irptc/sids/QECDSIDS/100005.pdf
(2) In the 13-week repeated-dose inhalation study in B6C3F1 mice described above, the gonadal
organs were evaluated in mice exposed to PNCB at 0, 6, 12, and 24 ppm. Treatment-related
effects on reproductive parameters included significantly increased estrus cycle length in females
"3
exposed to 24 ppm (153.6 mg/m /day). See human health data at
http://www.chem.unep.ch/irptc/sids/QECDSIDS/100005.pdf
(3) In a two-generation reproduction toxicity study, Sprague-Dawley rats (15 males/dose; 30
females/dose) were administered PNCB in corn oil via gavage at 0, 0.1, 0.7 or 5.0 mg/kg-day 7
days/week for 14 weeks prior to being mated. Treatment continued throughout mating, gestation
and lactation for a total of 167 days in the F0 generation. F1 rats (15 males/dose; 30
females/dose) were administered PNCB via gavage at identical concentrations for 7 days/week
for 18 weeks during a pre-mating growth period and then throughout mating, gestation, lactation
and a post-weaning period for a total of 219 days. F2 rats were sacrificed at postnatal day 21.
Mortalities in the F0 generation were 1, 0, 3, and 2 at 0, 0.1, 0.7 and 5.0 mg/kg-day, respectively;
all females and mostly attributed to gavage error. Hematology, including measurement of
methemoglobin levels, was not performed. Treatment did not have a statistically significant
effect on mean body weights and weight gains, male or female fertility indices, pregnancy rate,
length of gestation, the number of live pups at birth and pup weights during lactation. F0 males
treated with 5.0 mg/kg-day exhibited histological changes in the testes (2/15; bilateral
degeneration/atrophy of the epithelium and 1/15, bilateral maturation arrest of the germinal
epithelium). These same males, which did not mate, also exhibited oligospermia. Males treated
with 0.1 or 0.7 mg/kg-day were not examined for this endpoint. F0 females exhibited slightly
lower body weights at all treatment levels (not statistically significant). This finding was not
considered treatment-related due to an absence of a dose-related effect. Litter survival indices
were slightly lower than control at > 0.7 mg/kg-day. Pup survival index was significantly
reduced at 5.0 mg/kg-day, because 2 females treated with 5.0 mg/kg-day experienced complete
pup mortality within their litters. No external malformation or histopathological changes were
observed in dead pups recovered either at birth or during lactation in any treatment group. At 0.1
mg/kg-day, 1 dead pup recovered at birth had no tail. Mortality rates in the F1 adult rats at 0,
0.1, 0.7 and 5.0 mg/kg-day were: males, 6.7%, 13.3%, 33.3%, and 6.7% and in females 3.3%,
3.3%), 3.3%), and 3.3%>, respectively. Some deaths were attributed to gavage error.
Administration of PNCB in the F1 generation had no effect on mean body weights and gains,
food consumption during mating, gestation and lactation, litter and pup survival indices and pup
weights. Treatment-related effects observed in the F1 generation included an increase in
extramedullary hematopoiesis and brown pigmentation of reticuloendothelial cells in rats from
all groups; these effects were more pronounced in males and females at 5.0 mg/kg-day. F1
females at all dose levels exhibited a slightly lower mating index than the control group, but this
was not considered treatment-related due to the absence of a dose-response relationship. No
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evidence of toxicity or histological change was observed in F2 pups. Additional details are from
TSCATS (OTS0557087 and OTS0526382). See human health data at
http://www.chem.unep.ch/irptc/sids/QECDSIDS/100005.pdf
LOAEL (systemic toxicity) = 0.1 mg/kg-day (based on histopathological effects in the spleen
ofFl adults)
NOAEL (systemic toxicity) = Not established
NOAEL (reproductive toxicity) = could not be established (testes from control an high dose
groups only, were evaluated for histopathology; high dose group showed bilateral
degeneration/atrophy of epithelium of the testes and oligospermia)
(4) In an NTP, reproductive assessment by continuous breeding (RACB) study, Swiss CD-I mice
(20 breeding pairs/dose, treated; 40 breeding pairs, controls) were administered PNCB in corn
oil, via gavage at 0, 62.5, 125, or 250 mg/kg-day from 1 week prior to cohabitation, through 14
weeks of cohabitation for the F0 generation. The dams were allowed to nurse the F1 generation
and these animals were weaned at 21 days. The F1 generation was then exposed to the test
substance at 0 or 250 mg/kg-day from 1 week prior to cohabitation, through 14 weeks of
cohabitation. For the F0 generation, there were no treatment-related mortalities. Water
consumption was significantly reduced in males and females at 250 mg/kg-day. No other
changes in clinical signs were observed. Reduced pup weight and reduced pup weight gain were
observed in litters from dams treated with > 125 mg/kg. A decrease in pup viability during
nursing in animals from dams treated with 250 mg/kg was observed. There was also a reduction
in the number of pups born alive at the final litter. In the F1 generation, cyanosis was observed in
some females and many were noted to have enlarged spleens at necropsy (data not provided).
Adjusted liver weight was increased in males and females at necropsy . F1 males also exhibited
decreased seminal vesicle weight and F1 females exhibited increased estrous cycle length as
compared to controls. Pup viability was reduced and decreased adjusted pup weight was
observed at 250 mg/kg (only dose tested for this generation) in the offspring of F1 animals
(Chapin and Sloane, 1996; Chapin, 1997b). See human health data at
http://www.chem.unep.ch/irptc/sids/QECDSIDS/100005.pdf
LOAEL (reproductive and systemic toxicity) = 250 mg/kg-day (based on organ weight
changes and decreased fertility)
NOAEL (reproductive and systemic toxicity) = 125 mg/kg-day
Developmental Toxicity
ONCB (CASRN 88- 73-3)
In a prenatal developmental toxicity study, Sprague-Dawley rats (25 mated females/group) were
administered ONCB in corn oil, via gavage at 0, 25, 75 or 150 mg/kg-day on days 6 - 15 of
gestation. Observations were made on body weights, food consumption, number of live fetuses,
dead fetuses, early resorptions, implantations, corpora lutea, pre- and post- implantation loss,
pregnancy rate and the proportion of litters with abnormal external and skeletal fetal findings.
No clinical signs or post-mortem findings were observed in animals treated with 25 mg/kg-day.
Six females from the 150 mg/kg-day group died by gestation day 14 (four pregnant and two non-
pregnant) and their post-mortem findings did not indicate a specific cause of death. Due to
severe toxicity and high mortality, surviving females in the 150 mg/kg-day group were sacrificed
without necropsy examination prior to the date of scheduled sacrifice. At 75 mg/kg-day, dams
exhibited an increase in alopecia and urinary staining. At 150 mg/kg-day, dams exhibited
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urinary staining, pale extremities or eyes/cold to touch, alopecia, piloerection and
staining/encrustations on the face or forelimbs. There was a difference in maternal body weight
gain between dams treated with 75 mg/kg-day and controls that was accompanied by a decrease
in food consumption for gestation days 6-10. Most animals in the 150 mg/kg-day treatment
group exhibited body weight loss during gestation days 6-10, and all of the animals in this
group exhibited substantial reduction in food consumption during days 6-10 when compared to
the control group mean. There were no significant differences in pregnancy rate or the mean
number of live fetuses, dead fetuses, late resorptions, total implantations, corpora lutea or
preimplantation loss in the 25 and 75 mg/kg-day dose groups when compared to the control
group. A statistically significant increase in early resorptions and corresponding
postimplantation loss was exhibited in the 75 mg/kg-day treated group. Group mean fetal body
weight and sex distribution in the 25 and 75 mg/kg-day groups were comparable to the control
group. The total number of litters exhibiting external and skeletal malformations at 25 and 75
mg/kg-day dose groups was comparable to the control group. Increased incidences of skeletal
variations were observed in the 25 and 75 mg/kg-day group. Due to the high level of toxicity
observed at 150 mg/kg-day dose level, a separate test was conducted a year later with dosing at 0
or 100 mg/kg-day. This test group showed maternal toxicity as body weight loss for days 6-10
and reduced food consumption for days 6-16. The statistically significant increases in
resorptions, post-implantation losses and variations noted at 75 mg/kg-day in the first study,
were not observed in the 100 mg/kg-day test group. Additional details are from TSCATS
(OTS0524332). See human health data at
http ://www. chem .unep. ch/irptc/sids/OECD SID S/CHLORONITROB .pdf.
LOAEL (maternal toxicity) = 75 mg/kg-day (based on decreased maternal body weight gain)
NOAEL (maternal toxicity) = 25 mg/kg-day
NOAEL (developmental toxicity) = could not be determined
PNCB (CASRN100-00-5)
(1) In a prenatal developmental toxicity study, Sprague-Dawley rats (24 mated females/group)
were administered PNCB in corn oil via gavage at 5, 15 or 45 mg/kg-day on gestation days 6 -
19. No treatment related mortalities were observed. Terminal body weight and body weight
gain was significantly reduced at 45 mg/kg-day. Mean spleen weights were significantly
increased in all treated groups, compared to controls and mean spleen-to-body weight ratios were
significantly increased in dams treated with >15 mg/kg-day. The mean number of resorptions
was significantly higher in the 45 mg/kg-day treatment group and the number of live fetuses was
significantly decreased in this same dose group. The mean number of male and female fetus and
the respective mean weights were also decreased at 45 mg/kg-day. A significant increase in the
incidence of skeletal malformations was observed in fetuses from the 45 mg/kg-day group. No
other adverse effects were observed. See human health data at
http://www.chem.unep.ch/irptc/sids/QECDSIDS/100005.pdf
LOAEL (maternal toxicity) = 15 mg/kg-day (based on reduced body weight gain and
increased spleen weights)
NOAEL (maternal toxicity) = 5 mg/kg-day
LOAEL (developmental toxicity) = 45 mg/kg-day (based on increased resorptions, decreased
live births and decreased fetal weight and increased skeletal malformations)
NOAEL (developmental toxicity) =15 mg/kg-day
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(2) In a prenatal developmental toxicity study, New Zealand White rabbits (18 mated
females/group) were administered PNCB via gavage at 0, 5, 15 or 40 mg/kg-day on gestation
days 7 - 19. All surviving females were killed on day 30 of gestation and the fetuses were
examined for malformations. Mortality rates were 1/18, 1/18, 1/18, and 8/18 at 0, 5, 15 and 40
mg/kg-day, respectively. The 40 mg/kg-day treatment group was terminated on day 20 of
gestation due to the high mortality rate and the occurrence of spontaneous abortions in two
females. One female from the control group also underwent a spontaneous abortion. Clinical
signs included soft stool at> 5 mg/kg-day, anogenital staining at 15 mg/kg-day and grayish/pale
appearing eyes at 40 mg/kg-day. Premature delivery occurred in 3 does treated with 5 mg/kg-
day and in 1 doe at 15 mg/kg-day. In all other surviving dams, there were no significant changes
in reproductive parameters (mean number of implantations, resorptions and fetuses). There was
a slight increase in the incidence of fetal malformations at 5 and 15 mg/kg-day. There was some
variability in the sex distribution ratio between control and treated (5 and 15 mg/kg-day)
animals, but this was not considered to be treatment related. No treatment related effects were
observed on fetal weight or fetal ossification. Additional details are from TSCATS
(OTS0557074). See human health data at
http://www.chem.unep.ch/irptc/sids/QECDSIDS/100005.pdf
LOAEL (maternal toxicity) = 5 mg/kg-day (based on clinical signs and premature delivery)
NOAEL (maternal toxicity) = Not established
LOAEL (developmental toxicity) = 5 mg/kg-day (based on increased incidence of
malformations)
NOAEL (developmental toxicity) = Not established
(3) In a range-finding prenatal developmental toxicity study, New Zealand White rabbits (5
females/group) were administered PNCB via gavage in corn oil at 5, 15, 45, or 135 mg/kg-day
from gestation days 7-19. All surviving females were killed on day 30 of gestation. Mortality
rates were 0/5, 0/5, 0/5, 1/5, and 5/5 at 0, 5, 15, 45, or 135 mg/kg-day, respectively. No
treatment-related effects were observed in 5, 15, or 45 mg/kg-day test groups with respect to
maternal body weight, uterine implantations, fetal weight and external malformations in fetuses.
One hundred percent mortality at 135 mg/kg-day precluded examination of fetuses. Grayish
appearing eyes were observed at each dose level during the dosing and post-dosing periods.
Females that died during the study were observed to have tracheal hemorrhage and fluid-filled
lungs. Additional details are from TSCATS (OTS0557081). See human health data at
http://www.chem.unep.ch/irptc/sids/QECDSIDS/100005.pdf
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(4) In the two-generation reproduction study in Sprague-Dawley rats previously described, litter
survival indices for the F0 generation were slightly lower than control at > 0.7 mg/kg-day. The
pup survival index was significantly reduced for the F0 generation at 5.0 mg/kg-day, because 2
females treated with this dose experienced complete pup mortality within their litters. No
external malformation or histopathological changes were observed in dead pups recovered either
at birth or during lactation in any treatment group for the F0 generation. In the F1 generation, no
adverse effect was indicated in regard to gestaion length, parturition, litter size, litter survival,
mean pup weight or pup sex distribution. See human health data at
http://www.chem.unep.ch/irptc/sids/QECDSIDS/100005.pdf
LOAEL (maternal toxicity) = 0.1 mg/kg-day (based on histopathological effects in the spleen
ofFl adults)
NOAEL (maternal toxicity) = Not established
LOAEL (developmental toxicity) = 0.1 mg/kg-day (based on decreased pup survival)
NOAEL (developmental toxicity) = Not established
Genetic Toxicity — Gene Mutation
In vitro
ONCB (CASRN 88- 73-3)
(1) In an NTP study, Salmonella typhimurium strains TA98 and TA100 were exposed to ONCB
in DMSO at 0, 10, 33, 100, 133, 166, 250, 333, 666, 1000 or 1666 |ig/plate with and without
metabolic activation. Positive and negative controls were included and responded appropriately.
Toxicity was observed at 666 |ig/plate without metabolic activation. Precipitation was not
observed (NTP, 1993).
CASRN 88-73-3 was mutagenic in this assay.
(2) In an NTP study, Salmonella typhimurium strains TA98 and TA100 were exposed to ONCB
in DMSO at 0, 3, 10, 33, 66, 100, 166, 333 or 666 |ig/plate with and without metabolic
activation. Positive and negative controls were included and responded appropriately.
Cytotoxicity and precipitation were not observed.
CASRN 88-73-3 was mutagenic in this assay.
(3) In an NTP study, Salmonella typhimurium strains TA98 and TA100 were exposed to ONCB
in DMSO at 0, 3.3, 10, 33, 100, 200 or 300 |ig/plate with and without metabolic activation.
Positive and negative controls were included and responded appropriately. Slight toxicity was
observed at 333 |ig/plate. Precipitation was not observed.
CASRN 88-73-3 was mutagenic in this assay.
(4) In an NTP study, Salmonella typhimurium strains TA98, TA100, TA1535 and TA1537 were
exposed to ONCB in DMSO at concentrations of 6 - 1000 |ig/plate with and without metabolic
activation. Positive and negative controls were included and responded appropriately. Toxicity
was observed at 600 and 1000 |ig/plate. Precipitation was not observed.
CASRN 88-73-3 was mutagenic in this assay.
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(5) In an NTP study, Salmonella typhimurium strains TA98, TA100, TA1535 and TA1537 were
exposed to ONCB in DMSO at 0, 10, 33.3, 100, 333.3, 1000 |ig/plate with and without metabolic
activation. Positive and negative controls were included and responded appropriately. Toxicity
was observed at 1000 |ig/plate. Precipitation was not observed.
CASRN 88-73-3 was mutagenic in this assay.
MNCB(CASRN 121-73-3)
(1) In an NTP study, Salmonella typhimurium strains TA98, TA100, TA1535 and TA1537 were
exposed to MNCB in DMSO at 0, 3.3, 10, 33, 100, 200, 250, 333, 400, 500 or 666 |ig/plate with
and without metabolic activation. Positive and negative controls were included and responded
appropriately. Toxicity was observed at concentrations of 500 and 600 |ig/plate. Slight toxicity
was observed at concentrations as low as 250 |ig/plate. Precipitation was not observed.
CASRN 121-73-3 was inconclusive in this assay.
(2) Salmonella typhimurium strains TA98, TA100, TA1535, TA1537 and TA1538 were exposed
to MNCB in DMSO at 25.6, 51.2, 102.4, 204.8, 409.6, 819.2, 1638.4 or 3276.8 ng/plate with and
without metabolic activation. Positive and negative controls were included and responded
appropriately. All tests were performed in duplicate and repeated at least three times.
Cytotoxicity was observed at 3276.8 |ig/plate. No information was provided with respect to
observed precipitation.
CASRN 121-73-3 was not mutagenic in this assay.
PNCB (CASRN 100-00-5)
(1) Salmonella typhimurium strains TA98, TA100, TA1535 and TA1537 were exposed to
PNCB at 0.01, 0.04, 0.2, 0.3, 1, 1.3, 1.5, 3, 4 or 10 mg/plate with and without metabolic
activation. Solvent (DMSO) and positive controls were included, but response data were not
provided. Cytotoxicity was observed at the concentration of 3 mg/plate. A definitive positive
response was observed in TA1535 without metabolic activation, with some indication of a
marginally positive response with metabolic activation.
CASRN 100-00-5 was mutagenic in this assay.
(2) Salmonella typhimurium strains TA1535, TA1537 and TA1538 were exposed to PNCB at
concentrations < 1000 |ig/plate with metabolic activation and < 1500 |ig/plate without metabolic
activation. Negative, or solvent, and positive controls were included and responded
appropriately. No information was provided with regards to observations of cytotoxicity or
precipitation. Additional details are from TSCATS (OTS0557117).
CASRN 100-00-5 was not mutagenic in this assay.
(3) Salmonella typhimurium strains TA98, TA100, TA1535, TA1537 and TA1538 were exposed
to PNCB in DMSO at concentrations of 600, 1200, 1800, 2400 or 3000 |ig/plate with metabolic
activation and at 100, 200, 300, 400 or 500 |ig/plate without metabolic activation. Positive and
negative controls were included and responded appropriately. No information was provided with
regards to observations of cytotoxicity or precipitation. PNCB demonstrated weak mutagenicity
in strain TA1535 with metabolic activation. Additional details are from TSCATS
(OTS0557119).
CASRN 100-00-5 was mutagenic in this assay.
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(4) L5178Y mouse lymphoma cells were exposed to PNCB at 25, 60, 100, 150, 300, 402, 504 or
600 |ig/mL without metabolic activation and at 42, 77, 105, 140, 175, 203, 252 or 350 |ig/mL
with metabolic activation. In a repeat of the experiment with metabolic activation, mouse
lymphoma cells were exposed to PNCB at 21 |ig/mL, in addition to the previously used
concentrations. In a range-finding study, precipitation was observed at concentrations
> 2000 |ig/mL, A 1% DMSO solvent control and positive control (500 |ig/mL ethyl
methanesulfonate without activation; 6 |ag/m L with activation) were included and responded
appropriately. Cytotoxic concentrations were not identified; however, results of testing at the
concentration of 504 jag/m L without metabolic activation were not analyzed due to reduced cell
survival. The first and second experiments with metabolic activation did not include a statistical
analysis of results at concentrations >105 and 203 |ig/mL, respectively, due to reduced cell
survival. A positive response was observed with and without metabolic activation. Additional
details are from TSCATS (OTS0557075).
CASRN 100-00-5 was mutagenic in this assay.
(5) Chinese hamster ovary (CHO) cells were exposed to PNCB in DMSO at 100, 200, 300, 350
or 400 |ig/mL and 100, 300, 500, 700 or 900 |ig/mL with and without metabolic activation,
respectively. In a preliminary cytotoxicity test at 1000 |ag/m L that did not include a vehicle, no
cytotoxicity was observed without metabolic activation and 38.9% relative survival was
observed with metabolic activation. In a cytotoxicity test that included a DMSO vehicle, 8.2 and
61.9% survival values were obtained from exposure at 1000 |ig/mL with and without metabolic
activation, respectively. Precipitation was observed at > 300 jag/mL with metabolic activation
and at > 300 |ag/m L without metabolic activation. Positive (200 |ag/m L ethylmethanesulfonate
without activation; 100 |ag/m L dimethyl nitrosamine with activation), solvent (DMSO) and
untreated controls were included and responded appropriately. Additional details are from
TSCATS (OTS0557076).
CASRN 100-00-5 was not mutagenic in this assay.
(6) Salmonella typhimurium strains TA98, TA100, TA1535, TA1537 and TA1538 were exposed
to PNCB in DMSO at 100, 250, 500, 1000, 2500, 5000 or 10,000 |ig/plate with metabolic
activation and at 100, 250, 500, 1000 or 2500 |ig/plate without metabolic activation. Positive
and negative controls were included and responded appropriately. Cytotoxicity was observed in
all strains at 2500 |ig/plate without metabolic activation. In a separate trial for TA1535 in which
2500 |ig/plate was the highest dose tested without metabolic activation, cytotoxicity was
observed at 2500 |ig/plate. With metabolic activation, cytotoxicity was observed in TA98 at
10,000 |ig/plate. In one trial conducted with metabolic activation, cytotoxicity was observed in
TA1535 at concentrations of 500 and 1000 |ig/plate, but not at the highest tested concentration of
2500 |ig/plate. No information was provided with regards to precipitation. PNCB was weakly
mutagenic for strains TA1535 and TA100 with metabolic activation. Additional details are from
TSCATS (OTS0557122).
CASRN 100-00-5 was mutagenic in this assay.
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(7) Salmonella typhimurium strains TA98, TA100, TA1535, TA1537 and TA1538 were exposed
to PNCB in DMSO at 250, 500, 1000, 2500 or 5000 |ig/plate with metabolic activation and at
100, 250, 500, 750 or 1000 |ig/plate without metabolic activation. Positive and negative
(DMSO) controls were included and responded appropriately. Cytotoxicity was observed in
strain TA1538 at 2500 and 5000 |ig/plate with metabolic activation. No information was
provided with regards to observations of precipitate. PNCB was weakly mutagenic for strain
TA100 with metabolic activation and showed possible mutagenic activity in strain TA1535 with
metabolic activation. Additional details are from TSCATS (OTS0557123).
CASRN 100-00-5 was mutagenic in this assay.
(8) Salmonella typhimurium strains TA98, TA100, TA1535, TA1537 and TA1538 were exposed
to PNCB in DMSO at 250, 500, 1000, 2500, 5000 or 10,000 |ig/plate with metabolic activation
and at 100, 250, 500, 1000 or 2500 |ig/plate without metabolic activation. Cytotoxicity was
observed in strains TA98 and TA100 at 2500 |ig/plate without metabolic activation. In one of
two trials, cytotoxicity was observed in strain TA100 at 5000 |ig/plate with metabolic activation.
Positive and negative (DMSO) controls were included and responded appropriately. No
information was provided with respect to observations of precipitate. PNCB was mutagenic for
strain TA1535 with metabolic activation and weakly mutagenic for strain TA100 with metabolic
activation. Additional details are from TSCATS (OTS0557124).
CASRN 100-00-5 was mutagenic in this assay.
(9) Salmonella typhimurium strains TA98, TA100, TA1535 and TA1537 were exposed to PNCB
in DMSO at 100, 250, 500, 1000 or 2500 |ig/plate with metabolic activation and at 100, 250,
500, 750 or 1000 |ig/plate without metabolic activation. One trial conducted for strains TA1535
and TA100 included exposure concentrations of 50, 100, 450, 500, 1000, 1500 and 2500
|ig/plate; cytotoxicity was observed in both strains at the highest concentration. Positive and
negative DMSO controls were included and responded appropriately. No information was
provided with respect to observations of precipitate. PNCB was mutagenic in strains TA1535
and TA100 with metabolic activation. Additional details are from TSCATS (OTS0557125).
CASRN 100-00-5 was mutagenic in this assay.
(10) Salmonella typhimurium strains TA98, TA100, TA1535 and TA1537 were exposed to
PNCB in DMSO at 250, 500, 1000, 2500 or 5000 |ig/plate with metabolic activation and at 100,
250, 500, 1000 or 2500 |ig/plate without metabolic activation. One trial conducted for strains
TA1535 and TA100, included exposure concentrations 50, 100, 250, 500 and 1000 |ig/plate,
with and without metabolic activation. Positive and negative (DMSO) controls were included
and responded appropriately. Cytotoxicity was observed at 2500 |ig/plate without metabolic
activation. No information was provided with regards to observations of precipitate. PNCB was
mutagenic for strains TA1535 and TA100 with and without metabolic activation. Additional
details are from TSCATS (OTS0557126).
CASRN 100-00-5 was mutagenic in this assay.
(11) CHO cells were exposed to PNCB in acetone at concentrations of 1.59 - 2.86 mM with and
without metabolic activation. Positive and negative controls were included and responded
appropriately. Toxicity was observed at higher concentrations (concentrations not specified).
The lower concentrations tested with and without activation were soluble in acetone and did not
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precipitate when added to the treatment medium. The higher concentrations tested with and
without activation were soluble in acetone and precipitated when added to the treatment medium.
Additional details are from TSCATS (OTS0557127).
CASRN 100-00-5 was not mutagenic in this assay.
(12) In an NTP study, Salmonella typhimurium strains TA98, TA100, TA1535 and TA1537 were
exposed to PNCB in DMSO at concentrations of 30 - 3000 |ig/plate with and without metabolic
activation. Positive and negative controls were included and responded appropriately. Slight
toxicity was observed at concentrations as low as 500 |ig/plate. Precipitation was not observed.
CASRN 100-00-5 was mutagenic in this assay.
(13) In a study conducted by NTP, Salmonella typhimurium strains TA98, TA100, TA1535,
TA1537, NR3 and NR101 were exposed to PNCB in DMSO at concentrations of 1 - 10,000
|ig/plate with and without metabolic activation. Positive and negative controls were included
and responded appropriately. Toxicity was observed in tests conducted at concentrations of
3333.3 and 10,000 |ig/plate in strains NR101 and TA100, respectively. Slight toxicity was
observed at concentrations as low as 666.7 |ig/plate. Precipitation was not observed.
CASRN 100-00-5 was mutagenic in this assay.
Genetic Toxicity — Chromosomal Aberrations
ONCB (CASRN 88- 73-3)
(1) In a NTP study, CHO cells were exposed to ONCB in DMSO at concentrations of 5 - 500
|ig/mL in the presence and absence of metabolic activation. Positive and negative controls were
included and responded appropriately. No information was provided with respect to
observations of cytotoxicity or precipitation. A weak positive response was observed in the
presence of metabolic activation.
CASRN 88-73-3 induced sister chromatid exchange in this assay.
(2) CHO cells were exposed to ONCB in DMSO at concentrations of 50 - 500 |ag/m L with and
without metabolic activation. Positive and negative controls were included. The negative
control response was appropriate and no information was provided with regards to the positive
control response. No information was provided with respect to observations of cytotoxicity or
precipitation.
CASRN 88-73-3 induced chromosomal aberrations in this assay.
(3) In a NTP study, CHO cells were exposed to ONCB in DMSO at concentrations of 5 - 500
|ig/mL with and without metabolic activation. Positive and negative controls were included and
responded appropriately. No information was provided with respect to observations of
cytotoxicity or precipitation. A positive response was observed without metabolic activation.
CASRN 88-73-3 induced sister chromatid exchange in this assay.
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(4) In a NTP study, CHO cells were exposed to ONCB in DMSO at concentrations of 16 - 500
|ig/mL with and without metabolic activation. Positive and negative controls were included and
responded appropriately. No information was provided with respect to observations of
cytotoxicity or precipitation.
CASRN 88-73-3 did not induce chromosomal aberrations in this assay.
MNCB(CASRN 121-73-3)
(1) In a NTP study, CHO cells were exposed to MNCB in DMSO at concentrations of 5, 16 or
50 |ig/mL without metabolic activation and at 1.6, 5 or 160 |ag/m L with metabolic activation.
Cells were examined for evidence of sister chromatid exchange. Positive and negative controls
were included and responded appropriately. No information was provided with respect to
observations of cytotoxicity or precipitation. The assay was inconclusive in the absence of
metabolic activation and negative in the presence of metabolic activation.
CASRN 121-73-3 was inconclusive in this assay.
(2) In a NTP study, CHO cells were exposed to MNCB in DMSO at concentrations of 50, 160 or
500 |ig/mL with and without metabolic activation. Positive and negative controls were included
and responded appropriately. No information was provided with respect to observations of
cytotoxicity or precipitation.
CASRN 121-73-3 did not induce chromosomal aberrations in this assay.
In vivo
PNCB (CASRN 100-00-5)
(1) Sprague-Dawley rats (5/sex/time period) were administered PNCB in corn oil via gavage at
doses of 30, 100 or 300 mg/kg-bw and sacrificed at 6, 12 or 24 hours after treatment. Positive
(cyclophosphamide) and negative (corn oil) controls were included but details of their responses
were not provided. Treatment-related effects included cyanosis in the mid- and high-dose groups
as well as weight loss in the high-dose group only. Additional details are fromTSCATS
(OTS0557083).
CASRN 100-00-5 did not induce chromosomal aberrations in this assay.
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Genetic Toxicity - Other
In vitro
PNCB (CASRN100-00-5)
(1) Rat hepatocytes in primary culture were exposed to PNCB in DMSO at 0, 3.33, 10.0, 33.3,
100, 333.3, 1000, 3333.33 or 10,000 |ig/well. Cytotoxicity was observed at concentrations >
1000 |ig/well. Positive (2-acetoamidofluorene), solvent (DMSO) and untreated controls were
included and responded appropriately. PNCB did not produce a mean grain count of five or
greater than the vehicle control mean grain count (2.5) at any level of concentration. It was
noted that the high, variable cytoplasmic counts observed for this assay may lead to false
negative or false positive results. Additional details are from TSCATS (OTS0557077).
CASRN 100-00-5 did not induce unscheduled DNA synthesis in this assay.
(2) Rat hepatocytes in primary culture were exposed to PNCB in acetone at 0.1, 0.5, 1, 5, 10, 50,
75, 100 or 500 |ig/mL. Cytotoxicity was observed at concentrations of 100 and 500 |ig/mL.
Precipitate was observed at 500 jag/m L in a solubility test, but was not observed when tested in
the preliminary experiment. Positive (2-acetylaminofluorene), solvent (acetone) and untreated
controls were included and responded appropriately. Additional details are from TSCATS
(OTS0557078).
CASRN 100-00-5 did not induce unscheduled DNA synthesis in this assay.
Additional Information
Skin Irritation
ONCB (CASRN 88- 73-3)
Six rabbits (strain and sex not specified) were administered 0.5 mL of a 10% solution of ONCB
in sesame oil to intact, abraded skin with semiocclusion, for 24 hours. Erythema was not
observed at 48 or 72 hours.
CASRN 88-73-3 was not irritating to rabbit skin in this study.
PNCB (CASRN 100-00-5)
(1) Six rabbits (strain and sex not specified) were administered PNCB in water, to shaved intact
skin at 500 mg for 24 hours with occlusion. The test substance was slightly irritating.
CASRN 100-00-5 was slightly irritating to rabbit skin in this study.
(2) Six rabbits (strain and sex not specified) were administered PNCB undissolved, to shaved
intact skin at 500 mg for 24 hours with occlusion. The test substance was not irritating.
CASRN 100-00-5 was not irritating to rabbit skin in this study.
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Eye Irritation
ONCB (CASRN 88- 73-3)
(2) Six rabbits (strain and sex not specified) were instilled with 100 mg undissolved ONCB into
1 eye of each rabbit. Observations were made at 1, 7, and 24 hours after application. The test
substance was slightly irritating.
CASRN 88-73-3 was slightly irritating to rabbit eyes in this study.
PNCB (CASRN 100-00-5)
(1) Albino rabbits (two males; strain not specified) were given an ocular administration of PNCB
(purity 99.5%) as 10 mg of solid test material in the right conjunctival sac. One treated eye was
washed after 20 seconds while the other remained unwashed. Observations of the cornea, iris
and conjunctiva were made at 1 and 4 hours, and at 1, 2 and 3 days. A small area of transient,
slight corneal cloudiness and no iritic or conjunctival effects were observed in the washed eye.
The washed eye was normal 4 hours after treatment. No corneal, iritic or conjunctival effects
were observed in the unwashed rabbit eye. Additional details are from TSCATS (OTS0557116).
CASRN 100-00-5 was not irritating to rabbit eyes in this study.
(2) Rabbits (strain, sex not specified; n=6) were instilled with 100 mg undissolved PNCB into 1
eye of each rabbit. Observations were made at 1, 7, 24, 48, 72 hours and day 8 after application.
The test substance was not irritating.
CASRN 100-00-5 was not irritating to rabbit eyes in this study.
(3) Rabbits (strain, sex not specified; n=6) were instilled with 0.1 mL of polymorphic PNCB into
1 eye of each rabbit. The eyes were not rinsed. Observations were made at 24, 48, 72 hours and
day 8 after applications. The test substance was not irritating.
CASRN 100-00-5 was not irritating to rabbit eyes in this study.
Carcinogenicity
PNCB (CASRN 100-00-5)
In the 24-month, gavage study in rats previously described, a higher incidence of interstitial cell
tumors of the testes in treated animals were observed compared to control animals (1.7%, 6.8%,
8.3%), and 10%> at 0, 0.1, 0.7, and 5.0 mg/kg-day, respectively). The authors believe this is due to
an unusually low incidence of these tumors in the control group rather than an oncogenic effect
of PNCB. Additional details are from TSCATS (OTS0557088).
CASRN 100-00-5 was carcinogenic to rats in this study.
Conclusion: The acute oral, inhalation and dermal toxicity of the chloronitrobenzenes in rats is
moderate. The acute dermal toxicity of PNCB in rabbits is low. In a 13-week, inhalation
repeated-dose toxicity study, rats exposed to ONCB exhibited an increase in methemoglobin
levels and hyperplasia and hypertrophy of the respiratory epithelium at 0.007 mg/L/day; the
NOAEC was not established. In a 13-week, inhalation repeated-dose toxicity study, mice
exposed to ONCB exhibited increased liver and spleen weights and hepatocytomegaly in the
liver at 0.0576 mg/L/day; the NOAEC is 0.0288 mg/L/day. In a 5-week dietary study in mice,
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exposure to ONCB produced histopathological changes in the liver in males and females at 167
mg/kg-bw/day and 220 mg/kg-bw/day, respectively; the NOAELmaies is 16 mg/kg-bw/day and
the NOAELfemaies is 24 mg/kg-bw/day. In a 13-week, inhalation repeated-dose toxicity study,
mice exposed to PNCB exhibited organ weight changes and histopathology in the liver at 0.0768
mg/L/day; the NOAEC is 0.0384 mg/L/day. In a 24-month oral gavage study, rats exposed to
PNCB exhibited increased methemoglobin levels at 0.7 mg/kg-day; the NOAEL is 0.1 mg/kg-
day. In an NTP, reproductive assessment by continuous breeding (RACB) study (oral gavage),
mice exposed to ONCB, exhibited an increase in methemoglobin levels and organ weight
changes at 80 mg/kg-day. There were no treatment related effects on the reproductive
parameters examined. The NOAEL for systemic and reproductive toxicity is 160 mg/kg-day
(highest dose tested). In an NTP, reproductive assessment by continuous breeding (RACB)
study (oral gavage), female mice exposed to PNCB, exhibited decreased fertility and organ
weight changes at 250 mg/kg-day; the NOAEL for systemic and reproductive toxicity was 125
mg/kg-day. In a prenatal developmental toxicity study in rats exposed to ONCB via oral gavage,
pregnant dams exhibited decreased maternal weight gain and increased early resorptions and post
implantation losses at 75 mg/kg-day; the NOAEL for maternal toxicity is 25 mg/kg-day. In a
prenatal developmental toxicity study in rats exposed to PNCB via oral gavage, pregnant dams
exhibited reduced body weight gain and increased spleen weights at 15 mg/kg-day; the NOAEL
for maternal toxicity is 5 mg/kg-day. In the same study, increased resorptions and skeletal
malformations and decreased live births and fetal weight were observed in pups from the 45
mg/kg-day treatment group; the NOAEL for developmental toxicity is 15 mg/kg-day. In the
same study, kits exhibited an increased incidence in fetal malformations at 5 mg/kg-day. In the
two-generation reproductive toxicity study in rats previously described, there was a decrease in
pup survival at 0.1 mg/kg-day; the NOAEL for developmental toxicity was not established.
ONCB and PNCB are mutagenic in bacteria, induced sister chromatid exchange in mammalian
cells and induced chromosomal aberrations in mammalian cells in vitro. MNCB was not
mutagenic in bacteria and did not induce chromosomal aberrations in mammalian cells in vitro.
PNCB did not induce chromosomal aberrations in rats in vivo. PNCB did not induce
unscheduled DNA synthesis in mammalian cells in vitro. ONCB is not irritating to rabbit skin
and is irritating to rabbit eyes. PNCB is irritating to rabbit skin and not irritating to rabbit eyes.
PNCB was carcinogenic to rats in a 24-month oral gavage study.
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Tabic 4. Summary Tabic of the Screening Information Data Set as Submitted under the U.S. HPV Challenge
Program — Human Health Data
Endpoints
ONCB
(CASRN 88-73-3)
MNCB
(CASRN 121-73-3)
PNCB
(CASRN 100-00-5)
Acute Oral Toxicity
LDS0 (mg/kg)
219
400
294
Acute Inhalation Toxicity
LCS0 (mg/L)
3.2
No Data
3.2
(RA)
>16.1
Acute Dermal Toxicity
LDS0 (mg/kg)
400 (rabbits)
655 (rats)
No Data
400
(RA)
3020 (rabbits)
750 (rats)
Repeated-Dose Toxicity
NOAEL/LOAEL
Oral gavage (mg/kg-day)
No Data
(rat, 24 months)
LOAEL = 0.7
NOAEL = 0.1
(RA)
No Data
LOAEL = 0.7
NOAEL = 0.1
(RA)
(rat, 24 months)
LOAEL = 0.7
NOAEL = 0.1
Repeated-Dose Toxicity
NOAEL/LOAEL
Inhalation (mg/L/day)
(rat, 13 weeks)
LOAEL = 0.007
NOAEL = Not established
No Data
LOAEL = 0.007
NOAEL = Not established
(RA)
(rat, 13 weeks)
LOAEL = 0.0096
NOAEL = Not established
Reproductive Toxicity
NOAEL/LOAEL
Oral gavage(mg/kg-day)
Systemic Toxicity
Reproductive Toxicity
NOAEL = 160 (highest
dose tested)
NOAEL = 160 (highest
dose tested)
No Data
LOAEL = 0.1
Not established
(RACB study)
LOAEL = 125
NOAEL = 62.5
(RA)
(2-gen repro study)
LOAEL = 0.1
NOAEL = Not established
Not established
Developmental Toxicity
NOAEL/LOAEL
Oral gavage (mg/kg-day)
Maternal Toxicity
Developmental Toxicity
LOAEL = 75
NOAEL = 25
NOAEL = Could not be
determined
No Data
LOAEL = 15
NOAEL = 5
LOAEL = 45
NOAEL = 15
(RA)
(prenatal dev tox study,
rat)
LOAEL = 15
NOAEL = 5
LOAEL = 45
NOAEL = 15
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Table 4. Summary Table of the Screening Information Data Set as Submitted under the U.S. HPV Challenge
Program — Human Health Data
Endpoints
ONCB
(CASRN 88-73-3)
MNCB
(CASRN 121-73-3)
PNCB
(CASRN 100-00-5)
Genetic Toxicity - Gene
Mutation
In vitro
Positive
Negative
Positive
Genetic Toxicity -
Chromosomal
Aberrations
In vitro
Positive
Negative
No Data
Positive
(RA)
Genetic Toxicity -
Chromosomal
Aberrations
In vivo
No Data
Negative
(RA)
No Data
Negative
(RA)
Negative
Genetic Toxicity - Other
In vitro
Unscheduled DNA
Synthesis
No Data
Negative
(RA)
No Data
Negative
(RA)
Negative
Additional Information
Skin Irritation
Not irritating
—
Irritating
Eye Irritation
Irritating
—
Not irritating
Carcinogenicity
—
—
Positive (rat, gavage)
Measured data in bold text; (RA) = read-across; — indicates that endpoint was not assessed for this substance.
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4. Hazard to the Environment
A summary of aquatic toxicity data submitted for SIDS endpoints is provided in Table 5. The
table also indicates where data for tested category members are read-across (RA) to untested
members of the category.
Acute Toxicity to Fish
ONCB (CASRN 88- 73-3)
Common carp (Cyprinus carpio)
96-h LC50 = 25.5 mg/L
SIDS document:http://www.chem.unep.ch/irptc/sids/OECDSIDS/CHLORONITROB.pdf
MNCB(CASRN 121-73-3)
(1) Fathead minnow {Pimephalespromelas) were exposed to MNCB (98% pure) at average
measured concentrations of 3.15, 5.05, 7.25, 14.4 or 24.1 mg/L in replicate one and 2.8, 4.5,
10.0, 12.9 or 22.4 in replicate two under flow-through conditions for 96 hours. Unfiltered Lake
Superior water was used as the control. Water quality parameters measured during the test
included total hardness, alkalinity, dissolved oxygen, temperature and pH.
96-h LC50 = 18.8 mg/L
PNCB (CASRN 100-00-5)
(1) Rainbow trout (Oncorhynchus mykiss) were exposed to PNCB (99.21% purity) at nominal
concentrations of 1, 1.8, 3.2, 5.6 or 10 mg/L under static conditions for 96 hours. PNCB was
administered in an acetone solution. Solvent, untreated and positive (antimycin A) controls were
included. Tests were conducted in soft reconstituted deionized water, supplemented with 48 mg
NaHCCb, 30 mg CaSC>4, 30 mg MgSC>4 and 2 mg KCL per liter. Water quality parameters such
as pH, ammonia and temperature were measured and no significant changes were observed. A
NOEC value of 1.8 was calculated.
96-h LC50 = 6 mg/L
(2) Fathead minnow (Pimephales promelas) were exposed to PNCB in acetone at nominal
concentrations of 10, 18, 24, 32, 42, 56, 75 or 100 mg/L under static conditions for 96 hours.
Mortalities were observed at > 24 mg/L. Additional details are from TSCATS (OTS0557114).
96-h LC50 = 26.7 mg/L
(3) Zebrafish (Brachydanio rerio)
96 h LC50 = 14.36 mg/L
SIDS document: http://www.chem.unep.ch/irptc/sids/QECDSIDS/100005.pdf
(4) Golden orfe/Ide (Leuciscus idus)
96 h LC50 = 2 mg/L
SIDS document: http://www.chem.unep.ch/irptc/sids/QECDSIDS/100005.pdf
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Acute Toxicity to Aquatic Invertebrates
ONCB (CASRN 88- 73-3)
(1) Daphnia {Daphnia magna) were exposed to ONCB at nominal concentrations of 6.25, 12.5,
25, 50 or 100 mg/L under static conditions for 48 hours. Clean water and solvent (0.5 mg/L
DMF) controls were included. Water quality was measured for dissolved oxygen, pH, alkalinity,
hardness and temperature and no significant changes were observed in any parameter.
48-h EC50 = 41 mg/L
(2) Daphnia (D. magna)
24-h EC50= 12 mg/L
48-h EC50 = 23.9 mg/L
SIDS document: http://www.chem.unep.ch/irptc/sids/OECDSIDS/CHLORONITROB.pdf
(3) Daphnia (J). carinata)
48-h EC50=21.3 mg/L
PNCB (CASRN 100-00-5)
(1) Daphnia (Daphnia magna) were exposed to PNCB (purity > 99%) at nominal concentrations
of 6.25, 12.5, 25, 50 or 100 mg/L under static conditions for 48 hours. PNCB was administered
in dimethyl formamide (DMF). Water quality was measured to record dissolved oxygen, pH,
alkalinity, hardness and temperature. A NOEC value of 3.2 was calculated.
48-h EC50 = 10 mg/L
(2) Daphnia (Daphnia magna)
48 h-ECso = 2.7 mg/L
SIDS-document: http://www.chem.unep.ch/irptc/sids/QECDSIDS/100005.pdf
Toxicity to Aquatic Plants
ONCB (CASRN 88- 73-3)
(1) Green algae (Scenedesmus subspicatus) were exposed to ONCB at nominal concentrations
of 0.8 - 100 mg/L under static conditions for 48 hours. Small deviations from the standard study
design included a shorter study duration (48 vs. 72 hours) and limited information presented on
each test concentration at each measurement point. Purity of ONCB was not noted.
48-h EC50 (biomass) = 34 mg/L
48-h EC50 (growth rate) = 75 mg/L
(2) Green algae (Chlorella pyrenoidosa)
96 11-1.1,( 50 = 6.9 mg/L
SIDS document: http ://www. chem .unep. ch/irptc/ sids/OECD SID S/CHLORONITROB .pdf
(3) Green algae {Scenedesmus subspicatus)
48h-ErC5o =75 mg/L
SIDS document: http ://www. chem .unep. ch/irptc/ sids/OECD SID S/CHLORONITROB .pdf
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PNCB (CASRN100-00-5)
(1) Green algae (Scenedesmus subspicatus) were exposed to PNCB at nominal concentrations of
0.8 - 100 mg/L under static conditions for 48 hours. Small deviations from the standard study
design included a shorter study duration (48 vs. 72 hours) and limited information presented on
each test concentration at each measurement point. Purity of PNCB was not noted.
48-h EC50 (biomass) = 8 mg/L
48-h EC50 (growth rate) = 16 mg/L
(2) Green algae {Scenedesmus subspicatus)
48 h-ErC5o =16 mg/L
SIDS document: http://www.chem.unep.ch/irptc/sids/QECDSIDS/100005.pdf
(3) Green algae (Chlorella pyrenoidosa)
96h-LOEC (growth rate) = 4.9 mg/L
SIDS document: http://www.chem.unep.ch/irptc/sids/QECDSIDS/100005.pdf
14-d Toxicity to Fish
ONCB (CASRN 88- 73-3)
Guppies (Poecilia reticulata) were exposed to ONCB (99% pure) at unspecified concentrations
under semi-static conditions for 14 days. Measured concentrations of test solutions were at least
80% of nominal. Water quality parameters measured during the test included oxygen content,
temperature and pH. No information on the use of a control group was provided.
14-d LC50 = 30.03 mg/L
PNCB (CASRN 100-00-5)
Zebrafish (Brachydanio rerio) were exposed to PNCB for 14 days.
14-d NOEC = 1.53 mg/L
Chronic Toxicity to Fish
ONCB (CASRN 88- 73-3)
Fathead minnow (Pimephales promelas)
33 d-NOEC = 0.264 mg/L (larval endpoint)
SIDS document: http://www.chem.unep.ch/irptc/sids/OECDSIDS/CHLORONITROB.pdf
Chronic Toxicity to Aquatic Invertebrates
ONCB (CASRN 88- 73-3)
Daphnia (Daphnia magna)
21 d-NOEC = 3 mg/L (reproduction)
SIDS document: http://www.chem.unep.ch/irptc/sids/OECDSIDS/CHLORONITROB.pdf
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PNCB (CASRN100-00-5)
(1)Daphnia {Daphnia magna) were exposed to PNCB for 21 days.
21 d-LOEC =0.103 mg/L (reproduction)
(2) Daphnia (.Daphnia magna) were exposed to PNCB for 21 days.
21d-NOEC = 0.190 mg/L (reproduction)
Conclusion:
MNCB (CASRN 121-73-3)
The 96-hour LC50 value for fish exposed to MNCB is 18.8 mg/L. The 48-hour EC50 value for
aquatic invertebrates exposed to MNCB is based on read-across from the range of values from
ONCB and PNCB as 2.7 to 41 mg/L. The 96-hour EC50 value for aquatic plants exposed to
MNCB is based on read-across from ONCB and PNCB ranged values of 6.9 to 34 mg/L for
biomass and 4.9 to 16 mg/L for growth rate. The 14-d NOEC for fish exposed to MNCB is based
on read-across from ONCB and PNCB as a range of values from 1.53 to 30.03 mg/L. The 33-day
NOEC for fish exposed to MNCB is based on the read- across value from ONCB as 0.264 mg/L.
The 21-day LOEC for MNCB based on the reproduction endpoint is read-across from PNCB as
0.103. The NOEC for MNCB is based on the range of read-across values from PNCB and ONCB
as 0.190 to 3 mg/L
ONCB (CASRN 88- 73-3)
The 96-hour LC50 value for fish exposed to ONCB is 25.5 mg/L. The 48-hour EC50 value for
aquatic invertebrates exposed to ONCB ranges from 21.3 to 41 mg/L. The 96 hour EC50 for
aquatic plants exposed to ONCB ranges from 6.9 to 34 mg/L for biomass. The 14-day fish
NOEC for ONCB is 30.03 mg/L. The 33-day NOEC for fish exposed to ONCB is 0.264 mg/L.
The 21-day NOEC for aquatic invertebrates exposed to ONCB is 3 mg/L based on the
reproduction endpoint.
PNCB (CASRN 100-00-5)
The 96-hour LC50 value for fish exposed to PNCB ranges from 2 to 26.7 mg/L. The 48-hour
EC50 value for aquatic invertebrates exposed to PNCB ranges from 2.7 to 10 mg/L. The 96 hour
EC50 for aquatic plants exposed to PNCB was 8 mg/L for biomass and ranges from 4.9 to 16
mg/L for growth rate. The 14-day NOEC for fish exposed to PNCB is 1.53 mg/L. The 21-day
LOEC and NOEC for aquatic invertebrates exposed to PNCB are 0.103 mg/L and 0.190 mg/L,
respectively, based on the reproduction endpoint.
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Table 5. Summary of the Screening Information Data Set
as Submitted under the U.S. HPV Challenge Program - Aquatic Toxicity Data
Endpoints
ONCB
(CASRN 88-73-3)
MNCB
(CASRN 121-73-3)
PNCB
(CASRN 100-00-5)
Fish
96-h LCS0 (mg/L)
25.5
18.8
2 - 26.7
Aquatic Invertebrates
48-h ECS0 (mg/L)
21.3-41
No Data
2.7-41
(RA)
2.7-10
Aquatic Plants
96-h ECS0 (mg/L)
(biomass)
(growth rate)
6.9 - 34
75 (48-h)
No Data
6.9-34
4.9- 16
(RA)
8
4.9-16
Fish 14-d
NOEC (mg/L)
30.03
No Data
1.53 - 30.03
(RA)
1.53
Chronic Toxicity
to Fish
33 d-NOEC (mg/L)
0.264
No Data
0.264
(RA)
No Data
0.264
(RA)
Chronic Toxicity to
Aquatic Invertebrates
21-d LOEC (mg/L)
21-d NOEC (mg/L)
(reproduction)
3
No Data
0.103
0.190-3
(RA)
0.103
0.190
(Bold) = experimentally derived data (i.e.,
derived from modeling); (RA) = read-across;
substance, (-) = no data.
derived from testing); (e) = estimated data (i.e.,
- indicates that endpoint was not evaluated for this
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5. References
Chapin, R. E. 1997a. 2-Chloronitrobenzene. Environmental Health Perspectives 105.
Chapin, R. E. 1997b. 4-Chloronitrobenzene. Environmental Health Perspectives 105.
Chapin, R. E., and Sloane, R. A. 1996. Reproductive Assessment by Continuous Breeding:
Evolving Study Design and Summaries of Eighty-Eight Studies. Environmental Health
Perspectives 104.
Nair, R. S., Johannsen, F. R., Levinskas, G. J., and Terrill, J. B. 1986. Assessment of toxicity of
o-nitrochlorobenzene in rats following a 4-week inhalation exposure. Fundam Appl Toxicol 7,
609-614.
NTP (1993). NTP technical report on toxicity studies of 2-chloronitrobenzene and 4-
nitrochlorobenzene administered by inhalation to F344/N rats and B6C3F1 mice. National
Toxicology Program, Research Triangle Park, NC.
http://ntp.niehs.nih.gov/ntp/htdocs/ST rpts/TOX33.pdf.
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